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300 N. ZEEB ROAD, ANN ARBOR. Ml 48106
18 BEDFORD ROW, LONDON WC1R 4EJ, ENGLAND
8109045
BRILMAN, LEAH ANN MOORE
CHROMOSOME MORPHOLOGY IN CYNODON DACTYLON (L.) PERS.
The University of Arizona
University
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300 N. Zeeb Road, Ann Arbor, MI 48106
1981
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300 \ 2=== 3D.. ANN ARBOR Ml .18106 '3131 761-4700
CHROMOSOME MORPHOLOGY IN
CYNODON DACTYLON (L.) PERS.
by
Leah Ann Moore Brilman
A Dissertation Submitted to the Faculty of the
DEPARTMENT OF PLANT SCIENCES
In Partial Fulfillment of the Requirements
For the Degree of
DOCTOR OF PHILOSOPHY
WITH A MAJOR IN AGRONOMY AND PLANT GENETICS
In the Graduate College
THE UNIVERSITY OF ARIZONA
I
1 9 8 1
THE UNIVERSITY OF ARIZONA
GRADUATE COLLEGE
As members of the Final Examination Committee, we certify that we have read
the dissertation prepared by
Leah Ann Moore Brilman
entitled Chromosome Morphology in Cynodon dactylon (L.) Pers.
and recommend that it be accepted as fulfilling the dissertation requirement
for the Degree of
Doctor of Philosophy
.
t
/ -
Date
Dat«
i0
P
Date
^
(AJ
Final approval and acceptance of this dissertation is contingent upon the
candidate's submission of the final copy of the dissertation to the Graduate
College.
I hereby certify that I have read this dissertation prepared under my
direction and recommend that it be accepted as fulfilling the dissertation
requirement.
Dissertation Director
—t
/ /Date
^
/ 9'^/
STATEMENT BY AUTHOR
This dissertation has been submitted in partial
fulfillment of requirements for an advanced degree at The
University of Arizona and is deposited in the University
Library to be made available to borrowers under rules of
the Library.
Brief quotations from this dissertation are
allowable without special permission, provided that
accurate acknowledgment of source is made. Requests for
permission for extended quotation from or reproduction of
this manuscript in whole or in part may be granted by the
head of the major department or the Dean of the Graduate
College when in his judgment the proposed use of the
material is in the interests of scholarship. In all other
instances, however, permission must be obtained from the
author.
Signed : CfjLlh. d,
C/fflSA/tf
ACKNOWLEDGMENTS
I wish to extend thanks to my dissertation
committee, Drs. W.R. Kneebone, J.E. Endrizzi, W.P. Bemis,
K. Matsuda, and J.O. Anderson, for their assistance and
advice during my graduate studies. Special thanks to Dr.
W.R. Kneebone for serving as my major advisor and to Dr.
J.E. Endrizzi for giving me laboratory space.
I would like to thank my parents, Mr. and Mrs.
Charles F. Moore, for instilling in me a love of learning
and filling our house with love. The warmest thanks go to
my husband, Howard, whose loving support and constant
encouragement made all this possible.
iii
TABLE OF CONTENTS
Page
LIST OF ILLUSTRATIONS
v
LIST OF TABLES
vii
ABSTRACT
viii
INTRODUCTION
1
LITERATURE REVIEW
3
MATERIALS AND METHODS
18
Pachytene Chromosome Morphology
Somatic Chromosome Morphology
Production of Polyhaploids
Method Is Twin Seedlings
Method 2: Anther Culture
Giemsa C-banding
RESULTS
18
20
20
20
21
22
24
Pachytene Morphology in Tetraploid C. dactylon....
Somatic Chromosome Morphology
Production of Polyhaploids
Giemsa C-banding
24
41
45
46
DISCUSSION
50
SUMMARY
57
APPENDIX 1i
PACHYTENE CHROMOSOME MEASUREMENTS FROM
NINE TETRAPLOID CLONES OF CYNODON
DACTYLON YAR. DACTYLON
LIST OF REFERENCES
59
78
iv
LIST OF ILLUSTRATIONS
Figure
1.
Page
Pachytene chromosome complement of R17P4 with
bivalents numbered, genome and centromere
indicated by arrow
25
Composite karyotype of tetraploid Cynodon
dactylon var. dactylon
27
Chromosome 1-D' from R7P5 with the terminal knob
missing from the long arm
27
Chromosome 1-D' from 9946a with two prominent
chromomere pairs near the centromere of the
long arm and a small chromomere one-third
distal
27
Chromosome 2-D' from 9946a missing the short arm
on one chromosome of the bivalent
27
Chromosome 4-D from R35P6 with the ends missing
from the long and short arms
34
Chromosome 4-D from FB49 lacking the knob on the
short arm
34
Chromosome 4-D from R7P5 with one chromosome of
bivalent pair missing and one small
34
Chromosome 4-D from R2P5 with the end of the
short arm single stranded and not attached
to the NOR
34
10.
Chromosome 4-D from B296 not attached to the NOR
Jk
11.
Chromosome 5-D' from R7P5 with part of the short
arm missing at the end
34
Pachytene spreads from Tifgreen showing
bivalents and univalents
40
Accessory chromosomes from T32 with eight B
chromosomes, including a B trivalent
40
2.
3*
4.
5.
6.
7.
8.
9.
12.
13.
v
vi
LIST OF ILLUSTRATIONS—Continued
Figure
14.
Page
Accessory chromosomes from T51 with a B
chromosome, two telo B chromosomes and a
fragment
40
15.
Somatic chromosomes with an idiogram below
42
16.
Somatic spreads from T21
47
l?.
Diplotene-diakinesis spreads from T21
47
18.
Pachytene spread from T21 showing univalents and
bivalents
48
Giemsa C-bands from Yakima
48
19.
LIST OF TABLES
Table
1.
2.
3.
Page
G. dactylon clones used in this study with
chromosome number, variety and origin as"
shown
19
Average mean values and ranges among measurements
for pachytene lengths for all bivalents from
nine tetraploid clones
28
Mean values and ranges for somatic chromosome
lengths and arm ratios from diploid (d) and
tetraploid (t) clones
^3
vii
ABSTRACT
The pachytene bivalents of the tetraploid Cynodon
dactylon var. dact.ylon could all be identified on the
basis of characteristic morphological features including
knobs, prominent chromomeres, chromosome length, long arm
length, short arm length, arm ratio and relative length.
The bivalents could be separated into two sets of
chromosomes, D and D', with the D genome being homologous
to that found in the diploids, C. dactylon vars. aridus
and afghanicus. The two sets are homoeologous as shown by
the occasional formation of quadrivalents and the
closeness of chromosome measurements, although chromomere
patterns and knobs differed. C. dactylon var. dactylon
therefore appears to be a segmental allotetraploid.
In prophase somatic spreads of var. dactylon two
sets of chromosomes could also be separated by
heterochromatic patterns. In both the diploids, vars.
aridus and afghanicus, and the tetraploid the somatic
chromosomes could be homologised with their corresponding
pachytene bivalents. The chromosomes were of different
relative lengths during the two phases.
A polyhaploid obtained from a twin seedling of
C. dactylon var. dactylon showed both univalents and
viii
bivalents in a diplotene spread, supporting a segmental
allopolyploid origin of var. dactylon. This plant
reverted back to a tetraploid state before further
studies could be made.
Giemsa C-banding, using barium hydroxide at room
temperature for 50 minutes and 2xSSC at 60°C for 30
minutes to pretreat the slides before staining, produced
telomeric bands in the position of the knobs seen in
standard acetocarmine squash.
INTRODUCTION
Bermudagrass, Gynodon dactylon var. dactylon
(2n=36), is a tetraploid species important throughout
the world as a turf, forage and weed. The diploid
species, C. dactylon var. aridus (2n=18) or giant
bermudagrass, has been introduced to Arizona and Hawaii
where it is used as a forage. Another variety of
interest is C. dactylon var. afghanicus (2n=18, 36)>
which has been suggested by Harlan, de Wet, et al. (1970)
to form an introgression product with var. dactylon.
Triploid hybrid bermudagrasses used for turf are crosses
of the diploid species C_. transvaalensis (2n=18) with
var. dactylon.
Genomic relationships in Cynodon have been
postulated on the basis of pairing at metaphase I of
meiosis. An autopolyploid origin of var. dactylon from
var. aridus with essentially no genomic differentiation
has been suggested by many researchers (Harlan, de Wet,
and Rawal 1970; Gupta and Srivastava 1970; Rawal and
Chedda I97I). A segmental allopolyploid origin has
also been suggested on the basis of quadrivalent numbers
in var. dactylon and trivalents in the triploid hybrid of
var. aridus x var. dactylon (Sengupta 1968; Tripathi,
Sachdeva, and Malik 1977)*
1
2
Ourecky (19&3) studied the pachytene chromosome
morphology of four cells from a diploid C. dactylon. A
detailed study of the pachytene chromosome morphology of
the diploid C. dactylon was done by Brilman (1978). Initial
observations in this paper suggested the tetraploid var.
dactylon possessed two sets of the same chromosomes.
Further observations need to be made to further elucidate
their relationship to the diploid set. Many researchers have
suggested that the low number of quadrivalents in the
tetraploid and trivalents in the triploid could be due to
low chiasma frequency (Forbes and Burton 1963; Gupta and
Srivastava I970). Pairing behavior and appearance of the
chromosomes at pachytene would be more indicative of actual
homology in the tetraploid and triploids.
Pachytene pairing in 2x haploids (polyhaploids) may
help decide if one or two different genomes are present in
the tetraploid. It may also show if structural changes such
as translocations exist between genomes.
Giemsa C-banding has been used in many plant
species to differentiate between chromosomes. Different
patterns of staining have been obtained with this method and
could help in differentiating genomes.
LITERATURE REVIEW
The genus Cynodon has recently been revised
using primarily morphological traits but grouping
genetically allied units as a single species (Harlan
et al. 1966; Clayton and Harlan 1969; Sengupta 1968;
Harlan and de Wet 1969; de Wet and Harlan 1970; Harlan,
de Wet, and Rawal 1970; Harlan, de Wet, et al. 1970;
Rawal and Harlan 1971). This resulted in the two grasses
in this study, bermudagrass and giant bermudagrass,
being classified as C. dactylon (L.) Pers. var. dactylon
(2n=36) and C. dactylon (L.) Pers. var. aridus Harlan
et de Wet (2n=l8) respectively. Also used in this study
was C. dactylon (L.) Pers. var. afghanicus Harlan et
de Wet (2n=l8). C. dactylon var. dactylon is further
divided into three races: tropical, temperate and
seleucidus. G. dactylon var. aridus is divided into two
races, a large robust one and a small slow growing one
found in Southern India and Ceylon (Harlan and de Wet
1969).
There have been conflicting reports on the basic
chromosome number in the genus Cynodon. Hurcombe (19^7)
reported a basic number of x=10 from root tips, with
C. bradleyi (2n=l8) (now classified as C. incompletus)
being of an aneuploid origin. Other reports of x=10
4
included C. dactylon with forty chromosomes (Tateoka
195*0 and biotypes of C. dactylon with thirty and forty
chromosomes (Rochecouste 1962). The basic number of x=9
was also reported by many researchers (Burton 19^7;
Brown 1950; Burton 1951; Forbes and Burton 1963; Ourecky
1963; Gould 1966). Forbes and Burton (1963) concluded
that the previous reports of x=10 were due to the
presence of "fragments" that were found to be satellites
at the ends of long secondary constrictions. Later
studies concluded that the basic number was x=9 and
that reports of x=10 were due to the presence of
B-chromosomes in some accessions of Cynodon (Hoff 19^7;
Sengupta 1968; Powell et al. 1968).
The cytogenetics of the genus has been studied
at metaphase I of meiosis to elucidate specific
relationships and to determine the relationship of
pairing behavior to the fertility of specific cultivars.
Hoff (1967) studied meiotic behavior in Arizona clones of
bermudagrass (2n=36), giant bermudagrass (2n=18) and
apparent natural triploid hybrids between them. Giant
bermudagrass always had nine bivalents. Bermudagrass had
averages that ranged from 1.13 to 2.07 quadrivalents
in different clones, with ^7% of all quadrivalents
associated with the nucleolus. The triploids* averages
ranged from 2.7^ to 3•67 trivalents per cell with a
maximum pairing of seven trivalents. Gupta and Srivastava
5
(1971) reported a triploid from a natural stand with two
to fourteen univalents and a mean of 6.10 univalents, two
to ten bivalents with a mean of 7.15 bivalents, and zero
to five trivalents with a mean of 2.20 trivalents. Hanna
and Burton (1977) studied four tetraploid cultivars of
bermudagrass in which pairing varied from five per cent
of the cells having eighteen bivalents in one cultivar
to sixty-three per cent of its cells with eighteen
bivalents in another cultivar. These cultivars varied
from .51 "to .26 quadrivalents per cell.
Forbes and Burton ( 1 9 6 3 ) reported a maximum of
eight trivalents per cell in triploid hybrids between
diploid and tetraploid C. dactylon and a maximum of nine
trivalents in Tiffine, a triploid hybrid between
C. transvaalensis (2n=18) and tetraploid C. dactylon
(2n=36). A maximum of three quadrivalents was observed in
teraploid C. dactylon. The high rate of trivalent
formation in the triploids suggested homology between the
genomes of diploid and teraploid C. dactylon and between
those of tetraploid C. dactylon and diploid
C. transvaalensis. The low quadrivalent formation in the
tetraploid might indicate limited homology of two
genomes or advanced diploidization of a doubled set. On
the other hand the genomes could be completely homologous,
but with low chiasmata frequency limiting formation of
quadrivalents.
6
Sengupta (1968) found that C. dactylon var.
dactylon showed fairly regular meiotic behavior with a
quadrivalent, occasionally associated with the nucleolus,
and some univalents. Triploid hybrids betv/een C. dactylon
(2n=36) and C. transvaalensis (2n=l8) showed primarily
twelve bivalents and three univalents or thirteen
bivalents and one univalent, occasionally eleven
bivalents with five univalents, and nine bivalents with
nine univalents. He postulated that there may be two
partially homologous genomes, D'D* for C. transvaalensis
and DDD'D* for C. dactylon, or there may be preferential
pairing of the two sets. On the basis of pairing in
hybrids he suggested that C. transvaalensis,
C. incompletus (2n=l8, rarely 36), C. aethiopicus
(2n=l8,36), and C. nlemfuensis (2n=18,36) are allied to
C. dactylon vars. aridus and afghanicus, with DD genome.
Harlan, de Wet, et al. (1970) reported that
hybrids between C. transvaalensis and C. dactylon var.
dactylon (2n=18) showed essentially perfect pairing
suggesting no genome differentiation. Hybrids between
C. dactylon var. dactylon (2n=36) and C. transvaalensis
(2n=l8) showed seven to thirteen univalents (mean of 9.3)
and seven to ten bivalents (mean of 8.8). Hybrids
between var. aridus and var. dactylon showed six to nine
univalents, seven to twelve bivalents, zero to two
trivalents and zero to two quadrivalents. Hoff ( 1 9 6 7 )
/
7
found in eight synthetically produced aridus (2n=l8) x
dactylon (2n=3^) triploids the average number of
trivalents per cell from each plant ranged from J.Ok to
3.67. The most common association was three trivalents,
six bivalents, and six univalents. The above researchers
concluded that there appears to be no significant genome
differentiation in the species studied.
In triploids collected from natural stands of
C. dactylon, Gupta and Srivastava (1970) found zero to
six trivalents per cell with the maximum number in three
different clones ranging from four to six. Univalents
varied from four to thirteen with an average of 6.8 to
8.1 in the three clones. They concluded that the genomes
in diploid and tetraploid C. dactylon were homologous.
They speculated that the failure of univalents to
associate in trivalents was due to the small size of the
chromosome which rarely formed more than two chiasmata.
Tripathi et al. (1977) observed eighteen bivalents
and rarely one quadrivalent in natural tetraploids and
three to eight quadrivalents in colchiploids from var.
aridus. They believe that the few quadrivalents in
natural tetraploids may be due to partial homology of the
genomes or translocations. The tetraploid may be a
segmental allopolyploid from two diploids with considerable
homology. They also found pairing in triploids of primarily
nine bivalents and nine univalents which suggested to
8
them autosyndetic pairing. The few trivalents found
suggest either heterogenomic pairing, translocations or
some homologous segments distributed among the two
genomes of the tetraploid.
Rawal and Chedda (1971) produced hybrids between
various diploids and tetraploids in Cynodon and concluded:
1) the genomes have considerable homology, 2) polyploids
are essentially autopolyploids and 3) chromosome pairing
is essentially autosyndetic and therefore preferential.
The small number of univalents in 2n hybrids and
univalents and multivalents in polyploid hybrids suggest
that genie and chromosomal changes have occured in the
original genome during the course of evolution.
Using evidence from cytology, morphology and
ecology Rawal and Chedda (1971) postulated that the genus
evolved from an ancestor which was 2n, non-rhizomatous,
and distributed from South and East Africa to the Middle
East and India. A diploid form evolved rhizomes and
invaded more arid regions of Africa, the Near East and
India in which C. dactylon var. aridus represents the
modern survivor (Rawal and Harlan 1971). This is the only
diploid in the genus with rhizomes and the only likely
source of C. dactylon var. dactylon (Harlan and de Wet
1969). They concluded that var. dactylon is an
autopolyploid and that the morphologically smaller forms
9
of var. aridus appear to be closer to var. dactylon than the
"giant" race. Plants of the seleucidus race of var.
dactylon, often selected for grazing and hay production,
are probably an introgression product involving a
tetraploid race of C. dactylon var. afghanicus and a
temperate race of var. dactylon (Harlan, de Wet, et al.
1970). They found C. incompletus (2n=18, rarely 36) and
C. aethiopicus (2n=18, 36) to be effectively separated
from the other taxa and C. nlemfuensis var. nlemfuensis
(2n=18, rarely 36) to be closely related to C. dactylon
vars. aridus and afghanicus. G. nlemfuensis and var. aridus
may interact genetically in Southern Ethiopia.
C. transvaalensis could be considered a variety of
C. dactylon except it is morphologically distinct and shows
no introgression with sympatric taxa. C. dactylon var.
elegans (2n=36) was also considered to be an autotetraploid
from diploid var. aridus.
Thomas and Murray (1978) suggested that tetraploid
C. dactylon may be made up of autotetraploid and
allotetraploid individuals and is a heterogenous
assemblage of individuals of completely different origin.
Another possibility is that there is a genetic mechanisim
that promotes bivalent formation in tetraploids. Given a
recessive gene in the tetraploid that promoted bivalent
10
pairing instead of quadrivalents (aaaa) and diploids with
and without these alleles (AA, Aa, aa), then pairing
found in triploids could be explained. A triploid with Aaa
would be trivalent forming and a triploid with aaa would
be bivalent and univalent forming.
Metaphase I of meiosis and quadrivalent formation
are not always indicative of genome relationships and may
not be a reliable criterion for determining whether a
species is an auto or an allopolyploid. Hossain (1976)
found that in a random mating population of rye, disomic
association was the predominant type, whereas in inbred
materials the chromosome association pattern is
predominantly tetrasomic. This suggested that autoploids
which form multivalents with high frequency at the time
of their origin during subsequent evolution may shift to
the bivalent characteristic of allopolyploids, a
phenomenon referred to as diploidization. Gottschalk
(1972) found that in a Solanum stenotomum and
"S. a.iuscence" hybrid pachytene chromosomes varied in
total length but euchromatic regions paired normally. The
length differences were restricted to heterochromatic
regions so loops formed. However, MI had normal bivalents,
indicating that after spiralization the length
differences had disappeared. He concluded that studying
bivalent formation at MI only means that pairing and a
11
certain amount of homology exists, but it does not mean
that there is complete homology between the chromosomes.
He suggests meiotic pachytene is better to elucidate
genome relationships because spiralization has just
started and all structural elements can be analyzed.
Ourecky (1963) studied pachytene in one diploid
clone of G. dactylon from Afghanistan, which was probably
var. afghanicus because Harlan, de Wet, and Rawal (1970)
determined that this variety comes from that area.
Ourecky, using the entire chromosome complement of four
different meiotic cells, characterized the nine bivalents
on the basis of (1) linear length, (2) relative length,
(3) arm ratio, (4) number of prominent chromomeres and
(5) presence of terminal knobs and nucleolus. He found
that the average linear length, relative length and
presence and abscence of terminal knobs were the most
useful criteria for determining specific chromosomes.
Based on these characteristics he numbered the bivalents
from 1, the smallest, to 9> the largest, although some
bivalents in individual spreads showed variation. He
stated that the number of terminal knobs varies with the
strain and ecotype.
Tripathi et al. (1977) observed chromosome
polymorphism in Cynodon species with some differences in
terminal knobs and relative lengths of knobs as compared
12
to Ourecky. They concluded that several chromosomal
biotypes exist within the two diploid and tetraploid
races of G. dact.ylon, although they showed no chromosomes
or measurements. They believe that the polymorphism can
easily be explained by considering the wide geographic
distribution of the species and the acclimatization to
widely diversified areas.
Brilman, Kneebone and Endrizzi (198I) studied the
pachytene chromosome morphology of five diploid
C. dactylon, three var. aridus and two hybrids between
var. afghanicus and var. aridus. In most cases the
chromosome measurements and morphological descriptions
were similar to those of Ourecky (1963). The arm ratios
and arm measurements for chromosomes 3 and ^ were
different. Chromosome 3 differences appeared to be due to
different centromere placement and designation of which
chromosome in the complement was represented. Chromosome ^
differences were due to the fact that in one cell Ourecky
studied this chromosome was very contracted. Chromosome 9
in his study also appeared to have more chromomeres than
those studied by Brilman et al. (1981).
In a study of pachytene chromosomes of Medicago
species closely related to M. sativa L., Gillies (1972)
found they had extremely similar chromosome arm ratios
and proportional lengths, suggesting one common
cytogenetic unit. Absolute lengths differed, possibly
due to differences in contraction, with achromatic
regions shortening faster. Gillies and Bingham (1971)
found that the chromosomes of diploid M. sativa were also
considerably longer than the chromosomes of tetraploid
alfalfa or 2x haploids. Haploids (2x) with pachytene
chromosome lengths similar to the tetraploid suggest that
greater chromosome contraction may be a property of the
tetraploid state still present in the 2x haploid state.
Ho and Kasha (197*0 found that M. sativa pachytene
chromosomes and their individual arms had different
contraction rates that could change relative chromosome
lengths and arm ratios. This was more noticeable in
submedian chromosomes, which had decreasing arm ratios,
and chromosomes with unequal distribution of heterochromatic and euchromatic regions.
Maguire (1962) found variability in lengths and
arm ratios of pachytene chromosomes of corn. This
variability was of two kinds, one contributing
approximately uniformly per unit length throughout the
genome and the other a characteristic of each chromosome
unrelated to length. She also studied the satellite
regions of chromosome 6 and found that it contained from
one to five chromomeres, with expression varying from
cell to cell within a sample (Maguire 1977). She
suggested that missing chromomeres may reflect failure
of condensation or differential aggregation during
condensation of subunits. Zecevic (197^) studied the
knobs on pachytene chromosomes of maize and found that
the number and position varied between populations.
A study of pachytene and somatic chromosomes of
tomato, Lycopersicum esculentum L., showed that both are
differentiated into proximal chromatic and distal
achromatic parts (Ramanna and Prakken 1967). All 12
bivalents and 12 somatic pairs could be identified and
each somatic pair could be homologised with its
corresponding bivalent. In comparing somatic and
pachytene contraction,
the chromatic parts in somatic
tissues were contracted by a factor of 4 to 5 over
pachytene contraction, whereas the achromatic parts were
contracted by a factor of 3 0 .
Hyde (1953) in studying the differentiated
chromosomes of PIantago ovata L. (2n=8) in meiosis and
mitosis found all chromosomes showed sharp and asymetric
differentiation at pachytene which corresponded in detail
to that in mitotic prophase. Both the meiotic and somatic
chromosomes were made up of deeply staining proximal
segments on either side of the centromere and much more
weakly chromatic distal segments.
15
De and Krishnan ( 1 9 6 6 ) studied the pachytene and
somatic chromosomes of Phaseolus mungo L. They found that
the chromosomes were of different relative lengths during
the two phases. The two Nor chromosomes were the second
and third longest at somatic metaphase but only seventh
and ninth longest during pachytene.
Polyhaploids have been utilized to show the
relationship between the parental genomes of polyploids.
Polyhaploids from autopolyploids often show complete
synapsis as reported in Medicago sativa L. (Gillies and
Bingham 1971) and Sorghum halepense x S. vulgare var.
sudanense (Duara and Stebbins 1952). In Phleum pratense
(2n=6x=42), the polyhaploid makes seven bivalents and seven
univalents at first meiotic metaphase, even though each
chromosome is triplicated (Nordenskiold 19^-5). Most
allopolyhaploids have a low level of bivalent formation,
which is generally assumed to be due to the association
of corresponding homoeologous chromosomes from the
component genomes (Huskins 1931)•
Anther culture has been used recently to obtain
haploids from rice, Oryza sativa L. (2n=24), (Chen and
Lin 1976; Chen 1977? Chen 1978); Western wheatgrass,
Agropyron smithii Rydb., green needlegrass, Stipa
viridula Trin., creeping foxtail, Alopecurus
arundinaceus Poir. (Lo, Chen and Ross 1980); big bluestem
16
Andropogon gerardi, and Indiangrass, Sorghastrum nutans
(Chen et al. 1979). In the above studies various media
were used including MS (Murishige and Skoog 1962), RM
(Linsmaier and Skoog 1965), B5 (Gamborg, Miller and Ojima
1968) and cereal (Potrykus, Harms and Lorz 1976). -Jarious
hormones were utilized including auxins, NAA and 2,4D,
and kinetin in different concentrations. Inflorescences,
young and unemerged, were often cultured directly on
the media.
Haploids have also been obtained by searching
ge r m i n a t i n g s e e d l i n g s f o r t w i n s e e d l i n g s . K r u s e ( I 9 8 O )
obtained monoploids and dihaploids of Beta vulgaris
using this technique. These plants have also been known
to spontaneously double as reported by Kruse (I98O) in
Beta vulgaris and Chase (1952) in maize.
Giemsa C-banding has been used extensively
recently to distinguish plant chromosomes in both meiotic
and mitotic material in Triticum aestivum, Aegilops
variabilis (Jewell 1979)• barley, Hordeum vulgare L.,
(Linde-Lauresen 1975» Noda and Kasha 1978a; Noda and Kasha
1978b), Avena species (Yen and Filion 1977). Anemone
blanda L. (Marks 197^)> Secale cereale L. (Singh and
Lelley 1975)> and Triticale (Merker 1973)* Various
methods have proved successful: (1) ASG technique (Merker
1973)? (2) HCl-NaOH technique (Noda and Kasha 1978a); and
(3) BSG technique (Yen and Filion 1977; Kimber et al. 1976;
Singh and Lelley 1975; Linde-Laursen 1975)- Both
centromeric and terminal/intercalary bands have been
observed using these methods.
MATERIALS AND METHODS
Pachytene Chromosome Morphology
Tetraploid clones of C. dactylon var. dact.ylon
were obtained for study from the crossing block
maintained by the University of Arizona at Casa Grande
Highway Farm, Tucson, Arizona or selected in the course
of this study. Clones utilized and their origins are
included in Table 1.
Young spikes were collected, before the boot had
split, between 9»00 A.M. and 12:00 P.M. for meiotic
analysis and fixed in 3 s 1 95c/° ethanol iglacial acetic acid.
These were stored in the refrigerator. The anthers were
removed from the spike and stained using standard
acetocarmine squash technique, with a trace of iron
added from an iron needle. Alternate heating over a hot
water bath and squashing was utilized to obtain well
spread and stained chromosomes. Cells with excellent
spreading and differentiation of all pachytene bivalents
were selected for analysis and photographed. The
chromosome measurements were made from the photomicro­
graphs obtained, utilizing a fine wire that was bent to
follow the curves of the chromosome pairs.
18
19
Table 1.
C. dactylon clones used in this study with
chromosome number, variety, and origin as shown.
Clone
2n
Variety
B*i42
18
afghanicus
B*J420P,
18
afghanicus x aridus
1-77-1
18
aridus
Yakima
18
aridus
I x Y
18
aridus
Tifgreen
27
Fults
36
C. transvaalensis x
dactylon
dactylon
B296
36
dactylon
CL1
36
dactylon
FB141
36
dactylon
R1?P4
36
dactylon
FB49
36
dactylon
R35P6
36
dactylon
R7P5
36
dactylon
R2P5
36
dactylon
99^6a
36
dactylon
T21
18, 36
dactylon
T32
36
dactylon
T51
36
dactylon
Origin
Herat, Afghanistan
(P.I. 223129)
Open-pollinated
offspring of B4^2
Unknown
Yakima, WA
A. Law, WSU, Pullman
1-77-1 x Yakima
Tifton, Georgia
Hybrid bermudagrass
Greeley, Colorado
Jess Fults, CSU
North Platte,
Nebraska
Arizona seed,
Curly leaf
Collected in Georgia
by G.C. Horn, FL
Open-pollinated
offspring of FB141
#3 green, Pensacola
Country Club, FL
Open-pollinated
offspring of FB^9
Open-pollinated
offspring of FB^9
Open-pollinated
offspring of Alicia
Athens, Greece
Twin seedling of
Arizona seed
Twin seedling of
Arizona seed
Twin seedling of
Arizona seed
20
The position and number of prominent chromomeres,
knobs and the NOR were used in the identification of each
chromosome. Other major characteristics used to identify
each chromosome were (1) linear length, (2) short arm
length, (3) long arm length, (4) arm ratio and (5)
relative length.
Somatic Chromosome Morphology
Mitotic material was obtained from root tips
produced on stolons of diploid and tetraploid C. dactylon
in water. The root tips were removed and placed in saturated
1-bromonapthalene for one hour to accumulate dividing
cells. They were then fixed in 3'1 95/° ethanol sglacial
acetic acid. Later they were hydrolyzed in IN HC1 at 60°C
for seven minutes or placed in a 1% cellulasesl^ pectinase
mixture for two hours. Standard acetocarmine squash
technique was then utilized and the procedure followed
as above.
Production of Polyhaploids
Method li
Twin Seedlings
Seeds of C. dactylon var. dactylon were soaked
in 10$ NaOGl solution for ten seconds. These were placed
in petri dishes lined with filter paper soaked in
water for germination. The germinating seeds were checked
for the presence of twin seedlnng. The two plants of
21
each twin were separated and transplanted. After growth,
stolons were placed in water and root tips obtained.
Somatic and meiotic chromosomes were studied as previously
described.
Method 2:
Anther Culture
Anther culture was attempted to obtain polyhaploids. Two media were tried each with varying
concentrations of indoleacetic acid (IAA-auxin source)
and kinatin. MS-2 medium (Murashige and Skoog 1962) as
modified by Fox (1963), in which the major salts were
reduced to half of their original stregths, and Nitsch
and Nitsch medium (Nitsch and Nitsch 1956), with 0.1 or
2.0 ppm IAA in combination with 0.2 or 2.0 ppm kinetin,
were prepared for callus induction. All media were
adjusted to pH 5*7 before autoclaving and solidified with
10 grams granulated agar/liter. Twenty milliliters were
dispensed into a 2.5 x 15 cm capped tube, autoclaved and
cooled at a slant. Inflorescences were picked before the
boot split, surface sterilized in 6% calcium hypochlorite
for five minutes, and rinsed for five minutes in sterile
distilled water. Three anthers from a single floret
were removed and cultured in a single vessel or forty
millimeter segments of inflorescence were cultured
in a vessel. They were maintained at 25°C, 16 hour
22
daylength with cool white fluorescent light of
approximately 10 W/m 2.
Giemsa C-banding
Slides of meiotic and mitotic chromosomes were
prepared as described above using either acetocarmine
or
acetic acid, with a phase contrast scope. The
coverslip was removed by floating it off upside down in
20:/o acetic acid. This was followed by 10 minutes in
1:1 glacial acetic acid:70^ ethanol, 10 minutes in
70°/°
ethanol, 10 minutes in 95^ ethanol and 10 minutes in
100^ ethanol, then air drying at least overnight.
Various C-band techniques were tried that have
proved successful in other species. (1) ASG technique:
The slides were incubated in 2xSSC (0.3M NaCl-0.3M
trisodium citrate adjusted to pH 7.0 with 0.1N HC1) at
60°C for 60 minutes, rinsed in distilled water for 5
minutes and air dried (Merker 1973)• (2) HCl-NaOH
technique: The slides were placed in IN HC1 at 60°C for
5 minutes, rinsed in tap water, dried at room temperature
for one-half day and immersed in 0.07N NaOH for 35
seconds (Noda and Kasha 1978a). (3) BSG technique: Slides
treated in a freshly made 5% barium hydroxide solution
for times ranging from 3 "to 60 minutes at room
temperature and at 60°c, rinsed in tap water for 10
minutes, incubated in 2xSSC for times ranging from JO to
60 minutes at 6o°C and rinsed in tap water for 10 minutes
(Yen and Filion 1977; Kimber et al. 1976; Singh and
Lelley 1975; Linde-Laursen 1975).
The slides prepared in the above methods were
then immersed in 2fo giemsa solution (Giemsa stock
solution in Sorensen phosphate buffer, pH 6.9) for
approximately 30 minutes, with periodic monitoring to
check staining intensity. Slides were rinsed, air dried
and mounted in Permount.
RESULTS
Pachytene Morphology in
Tetraploid C. dactvlon
In each spread the two homoeologous pachytene
bivalents for each of the chromosomes that have been
numbered from one to nine, could be distinguished on the
basis of prominent chromomeres and knobs. These were
designated D and D', as per Sengupta (1968), with D being
those bivalents reported in the diploid C. dact.ylon by
Brilman, Kneebone and Endrizzi (1981). The two sets were
designated D and D' rather than D and E due to the
occasional formation of quadrivalents and the closeness in
length of each pachytene pair. Variation in the number of
prominent chromomeres between the diploid D chromosomes
and the tetraploid D chromosomes can be attributed to the
greater contraction of chromosomes in all spreads of the
tetraploid. The longest chromosome of each pair in the
tetraploid just approaches the length of the shortest
pachytene bivalent in the diploid, which were entering
diakinesis.
Variation in linear measurement of chromosomes
occurred within and between clones due to stage of the
cell, chromosome contraction and degree of spreading.
Figure 1 shows a typical pachytene spread with the
24
25
chromosome strands, chromomeres and knobs showing
prominently. The schematic drawing shows the number and
genome designation of each chromosome and the position of
the centromere.
Figure 2 shows a composite karyotype of the entire
chromosome complement using the average measurements of
each chromosome as in Table 2. The position and number of
the prominent chromomeres and knobs are those usually
found for each chromosome. Variations in these
characteristics were found between some clones and these
will be described for those chromosomes involved. The
measurements for each clone are shown in Appendix 1.
Chromosome 1-D:
Usually this chromosome, with an
average length of 9-8 um, was the shortest chromosome in
the complement and had a relative length of 1.
Occasionally, in very contracted spreads, chromosome 3-D
or either chromosome ^ was shorter and was given a
relative length of 1. The average relative length of this
bivalent was 1.0^. The centromere was submedian, with an
arm ratio of 1.1*6. The long arm had a terminal knob and
the short arm was knobless. The long arm averaged 5«8 um,
with two prominent chromomere pairs next to the
centromere and a slightly less prominent pair one-third
the distance from the centromere. The short arm was
^.0 um, with a small chromomere pair next to the
Figure 3« Chromosome 1-D* from R?P5 with the terminal knob
missing from the long arm.
Figure 4. Chromosome 1-D' from 99^6a with two prominent
chromomere pairs near the centromere of the long arm and a
small chromomere one-third distal.
Figure 5« Chromosome 2-D' from 99^6a missing the short arm
on one chromosome of the bivalent.
27
10 1020 2o· 30 30' 40 4o· so so· so so' 7o 10· so so· 90 90'
SJJ
Figure 2~ Composite karyotype of tetraploid Cynodon
dactylon var. dactylon.
SJJ
Figure J.
Chromo some 1-D'
from R7P5
Figure 4.
Chromo some 1-D'
from 9946a
Figure 5.
Chromosome 2-D'
from 9946a
Table 2.
Average mean values and ranges among measurements for pachytene lengths
arm ratio for all bivalents from nine tetraploid clones.
Length (um)
Chromosome
Number
No.
Cells
1-D
(50)
1-D'
(50)
2-D
(50)
Whole
Chromosome
Arm ratio
Short arm
Long arm
9.8-1.9
(5-0-14.0)
4.0+0.9
(1.8-6.3)
5.8+1.2
(3.1-9.0)
9.7*1.9
(5.0-13.1)
3.9-0.8
(1.9-5.8)
5.8+1.3
(3.1-9.9)
(1.00-3.09)
11.2+2.0
(6.5-16.0)
4.4+0.8
(2.3-6.0)
6.9-1.5
(4.0-10.8)
(I.03-2.74)
Relative
Length
1.04+0.06
1.46+0.29
(1.02-2.30) (1.00-1.3 1 )
1.53-0.33
1.61+0.37
1.04+0.06
(1.00-1.24)
1.21+0.14
(1.01-1.59)
2-D'
(50)
11.3*2.1
(6.8-16.3)
4.4*1.1
(2.6-8.1)
6.9+1.4
(4.2-11.3)
1.21+0.13
I.6I+O.35
(1.01-2.40) (1.05-1.53)
3-D
(50)
11.7*2.3
(7.1-18.1)
2.9*^.6
(1.8-4.3)
8.7*2.0
(4.7-14.2)
1.26+0.19
3.06*0.59
(1.74-4.30) (1.00-1.77)
3-D"
(50)
11.9*2.4
(7.2-17.2)
3.8*D.9
(2.0-5.6)
8.1*2.0
(4.4-13.6)
1.27+0.16
2.26+0.65
(1.15-3.78) ( 1 . 0 0 - 1 . 6 5 )
4-D
(17)
11.9*2.7
(7.1-17.1)
5.0*1.3
(3.0-8.0)
6.9+1.7
(4.1-9.4)
1.31+0.22
1.39+0.28
(1.13-2.13) (1.00-1.97)
4-D*
(50)
12.7*2.6
(7.2-21.9)
5.4*1.1
(2.9-9.8)
7.4*1.7
(4.2-13.5)
1.37+0.21
1.39-^0.24
(1.00-2.08) (1.02-1.97)
5-D
(50)
12.7*2.6
(6.9-18.8)
4.8*1.2
(2.9-7.8)
7.9*1.9
(4.5-12.3)
1.36+0.14
1.72+0.52
(1.12-1.20)
(1.01-3.97)
5-D'
(50)
12.9*2.7
(7.8-19.9)
4.9*1.3
(2.6-8.1)
8.O-+L.9
(4.8-11.8)
1.69+0.48
1.39-^0.17
(1.02-2.87) (1.11-1.76)
Table 2.
Continued
Length (um)
Chromosome
Number
No.
Cells
6-D
(50)
6-D'
Whole
Chromosome
Relative
Length
Short arm
Long arm
14.4*3.2
(7.9-21.9)
5.5*1.8
(2.9-10.0)
8.9*1.9
(4.5-13.2)
1.71*0.54
1.54*0.19
(1.04-3.69) (1.14-1.91)
(50)
14.7*3.1
(8.6-22.1)
5.9*1.4
(3.1-9.9)
8.8*2.2
(4.6-15.9)
1.54*0.40
1.58*0.20
(0.99-2.56) (1.17-1.98)
7-D
(50)
1 6 . 5 - 3 .4
(10.4-25.1)
7.1*2.1
(3.0-12.0)
9.4-1.7
(6.1-13.9)
1.41*0.35
1.77*0.24
(0.91-2.53) (1.42-2.51)
7-D'
(50)
16.5-3.3
(10.0-23.4)
7.0-1.6
(3.0-10.5)
9.6-2.0
(5.8-13.8)
1.41-0.2 9
1.78*0.23
(0.95-2.50) (1.43-2.51)
8-D
(50)
18.8-3.9
(11.9-28.3)
7.6-1.6
(4.6-11.6)
11.2-2.7
(6.7-18.8)
2.02*0.28
1.49*0.33
(1.02-2.72) (1.54-2.79)
8-D'
(50)
19.2-4.4
(12.0-33.2)
8.0-2.0
(4.8-13.2)
11.2-2.9
(6.8-20.0)
1.44*0.34
2.05*0.29
(0.83-2.29) (1.62-3.03)
Arm ratio
9-D
(50)
22.2-5.1
(12.9-32.8)
8.9*2.0
(4.9-12.8)
13.3*3.6
(7.2-20.7)
2.38*0.40
1.50*0.31
(1.10-2.46) (1.70-3.76)
9-D'
(50)
22.5-5.3
(13.5-34.8)
8.9*2.0
(5.3-14.5)
13.6*3.7
(7.4-22.8)
2.41*0.40
1.53*0.30
(1.09-2.42) (1.78-4.00)
ro
\o
30
centromere, two prominent chromomere pairs slightly
distal to the former and two chromomere pairs near the
end, which sometimes appeared as a small knob. This made
the overall appearance of the short arm heterochromatineuchromatin- heterochromatin.
Chromosome 1-D':
This chromosome had the same
relative length, 1.04, as chromosome 1-D and an average
length of 9.7 urn. The centromere was submedian, with an
arm ratio of 1.53- The long arm had a terminal knob,
except in R7P5 where it is missing and a longer segment
may be missing (Figure 3)» The short arm was knobless.
The long arm was 5*8 um, with usually one (Figure 1) or
sometimes two (Figure 4) prominent chromomere pairs next
to the centromere and may (Figure 4) or may not (Figure 1)
have a small chromomere one-third the distance from the
centromere. The short arm was 3*9 um, with two prominent
chromomere pairs next to the centromere and two small
chromomere pairs near the end.
Chromosome 2-D:
This chromosome had an average
length of 11.2 um and a relative length of 1.21. The
centromere was submedian with an arm ratio of 1.6l. The
long arm had a terminal knob and the short arm had a small
knob that could be resolved occasionally into two
prominent chromomeres. The long arm was 6.9 um, with two
prominent chromomere pairs near the centromere and two
31
chromomere pairs slightly distal to the former. The
short arm was 4.4 um, with two prominent chromomere pairs
next to the centromere and two more pairs slightlydistal to the first two pairs.
Chromosome 2-D':
This chromosome had an average
length of 11.3 um and a relative length of 1,21. This
chromosome was less heterochromatic than chromosome 2-D.
The centromere was submedian, with the same arm ratio of
1.6l as above. The long arm had a terminal knob and the
short arm was knobless. The long arm was 6.9 um, with only
two small chromomere pairs near the centromere. The short
arm was 4.4 um, with a chromomere pair near the
centromere, two prominent chromomere pairs slightly
distal to the former, and a small chromomere pair on the
end. Clone 9946a is missing the short arm on one
chromosome of the bivalent (Figure 5)»
Chromosome 3-D:
The average length was 11.7 um
and the relative length was 1.26. The centromere was
submedian, with an arm ratio of 3.06. The long arm had a
terminal knob, and the short arm had four prominent
terminal pairs of chromomeres or knobs, which identify
this chromosome. The long arm was 8.7 um, with two
prominent chromomere pairs near the centromere and four
small chromomere pairs scattered up the arm. The short arm
was 2.9 urn, with the knobs making it appear primarily
heterochromatic.
Chromosome 3-D':
This bivalent had an average
length of 11.9 urn and a relative length of 1.2?. The
centromere was submedian, with an arm ratio of 2.26. Using
Student's t-test this arm ratio was significantly smaller
(t=6.44 P=0.001) than the arm ratio of chromosome 3-D- The
long arm had a knob. The long arm was 8.1 um, with two
chromomere pairs next to the centromere and four small
chromomere pairs scattered up the arm. The short arm was
3.8 um, which was significantly longer (t=5.88 P=0.001)
than the short arm of chromosome 3-D* It also had four
prominent chromomere pairs, but there was euchromatin
between each and the chromosome did not appear as
heterochromatic.
Chromosome ^-D:
This was the nucleolar organizing
(NOR) chromosome, the average length in the seventeen
spreads where it was normal was 11.9 um and the relative
length was 1.31« The centromere was submedian, with an arm
ratio of 1.39. Both arms have terminal knobs. The long arm
was 6.9 um, with two prominent chromomere pairs between
the centromere and the NOR, two prominent pairs distal to
the NOR, three or less spaced chromomere pairs and two
more pairs between these and the knob. In R35P6, a terminal
deficiency in the long arm involving one or both chromo­
somes was apparent by the presence of single stranded
33
sections. The arm was significantly shorter at 7.1 urn
(t=3.66 P=0.005) than the long arm in chromosome 4-D' of
this clone at 10.2 um (Figure 6, Appendix 1). The short
arm was 5»0 um, with a chromomere pair next to the
centromere and approximately three more pairs between the
former and the knob. In R35P6 the short arm was also
missing the end, making the arm at 4,6 um significantly
shorter (t=3.94 P=0.005) than the short arm in chromosome
4-D' of this clone at 7.0 um (Figure 6, Appendix 1). FB49,
the maternal parent of R35P6, lacked the knob on the short
arm of this chromosome, but it was not significantly
shorter at 4.5 um than chromosome 4-D' of this clone at
4.9 um (Figure 7. Appendix 1). In R7P5. another progeny of
FB49, it appeared one chromosome of the pair may be
missing and one may be small (Figure 8). R2P5 was single
stranded on the end of the short arm and was not attached
to the NOR in five spreads (Figure 9). In B296 in three
spreads the chromosome appeared normal, but it is was not
attached to the NOR (Figure 10).
Chromosome 4-D':
This chromosome had an average
length of 12.7 um and a relative length of 1.37. The
centromere was submedian, with an arm ratio of 1.39. This
chromosome had a terminal knob on the short arm only, as
opposed to both arms on chromosome 4-D. The long arm was
7.4 um, with two prominent chromomere pairs between the
Figure 6. Chromosome 4-D from R35P6 with the ends missing from
the long and short arms.
Figure 7« Chromosome 4-D from FB49 lacking the knob on the
short arm.
Figure 8. Chromosome 4-D from R?P5 with one chromosome
of bivalent pair missing and one small.
Figure 9« Chromosome ^--D from R2P5 with the end of the
short arm single stranded and not attached to the NOR.
Figure 10.
NOR.
Chromosome 4-D from B296 not attached to the
Figure 11. Chromosome 5-D' from R7P5 with part of the
short arm missing at the end.
34
Figure 6. Chromosome 4-D
from RJ5P6
Figure 8.
from R7P5
Chromosome 4-D
Figure 7. Chromosome
4-D from FB49
Figure 9.
from R2P5
Chromosome 4-D
Sp
Figure 10.
from B296
Chromosome 4-D
Figure 11. Chromosome
5-D' from R7P5
centromere and the NOR, two prominent chromomere pairs
distal to the NOR, three small pairs, and two chromomere
pairs between these and the chromomere pair on the end.
The short arm was
um, with approximately four
prominent chromomere pairs between the centromere and the
knob. The short arm appeared more heterochromatic than in
chromosome
D.
chromosome 5-Ds
This chromosome had an average
length of 12.7 um and a relative length of 1.J6. The
centromere was submedian, with an arm ratio of l.?l. The
long arm had a terminal knob and there are two
chromomeres or a knob on the end of the short arm. The
long arm was 7.9 um, with two prominent chromomere pairs
near the centromere and one chromomere pair more distal.
In 99^6a there were three prominent chromomere pairs
near the centromere, two small pairs and then one pair
more distal. The short arm was 4.8 um, with one prominent
chromomere pair, one chromomere pair, one prominent pair
and the prominent chromomeres or knob on the end. The
whole chromosome appeared more heterochromatic than
chromosome 5-D'» especially the short arm.
Chromosome 5-D':
The average length of this
chromosome was 12.9 um and the relative length was 1.39.
The centromere was submedian, with an arm ratio of I.69.
There was only a terminal knob on the long arm. The long
arm was 8.0 um, with two prominent chromomere pairs near
the centromere. The short arm was 4.9 um. with three
chromomere pairs near the centromere and two small pairs
more distal. In R7P5 a terminal segment of the short arm
appears to be missing (Figure 11).
Chromosome 6-D:
The chromosome had an average
length of 14.4 um and a relative length of 1.54. The
centromere was submedian, with an arm ratio of 1.71. The
long arm had a terminal knob and the short arm had a
terminal chromomere pair or small knob. The long arm was
8.9 um, with three prominent chromomere pairs near the
centromere and one additional pair in the mid region of
arm. The short arm was 5«5 um, with three prominent
chromomere pairs near the centromere and two additional
pairs near the terminal knob or chromomere.
Chromosome 6-D':
The average length of the
chromosome was 14.7 um and the relative length of 1.58.
The centromere was submedian, with an arm ratio of 1.54.
The long arm had a terminal knob, with no knob on the
short arm. The long arm was 8.8 um, with three prominent
chromomere pairs near the centromere. The short arm was
5»9 um, with three prominent chromomere pairs near the
centromere and two small pairs distal to these.
Chromosome 7-Ds
The chromosome had an average
length of 16.5 um and a relative length of 1.77. The
37
centromere was submedian, with an arm ratio of 1.41. The
long arm had a terminal knob, but the short arm lacked
one, unlike chromosome 7 in the diploid as reported by
Brilman et al. (1981). The long arm was 9.4 um, with one
prominent chromomere pair near the centromere and two more
pairs slightly distal. The short arm was 7.1 um, with two
prominent chromomeres near the centromere.
Chromosome 7-D':
The chromosome had an average
length of 16.5 um and a relative length of 1.78. The
centromere was submedian, with an arm ratio of 1.41. The
long arm had a terminal knob and the short arm had a
terminal prominent chromomere or small knob. The long arm
was 9-6 um, with three prominent chromomere pairs near the
centromere and three small pairs distributed along the arm.
The short arm was 7.0 um, with four prominent chromomere
pairs spaced along the arm. This chromosome was more
heterochromatic than chromosome 7-D or chromosome 7 in the
diploid.
Chromosome 8-D:
This chromosome had an average
length of 18.8 um and a relative length of 2.02. The
centromere was submedian, with an arm ratio of 1.49. The
long arm had a terminal knob and the short arm a terminal
chromomere pair or small knob. The long arm was 11.2 um,
with one prominent chromomere pair, three spaced pairs and
two small chromomere pairs in the distal one-third region,
38
a space, and two additional pairs. The short arm was 7.6
um, with three spaced prominent chromomere pairs and two
additional small pairs between these and the terminal
knob.
Chromosome 8-D's
The chromosome had an average
length of 19.2 um and a relative length of 2.05. The
centromere was submedian, with an arm ratio of 1.*j4. The
long arm had a terminal knob. The long arm was 11.2 um,
with two prominent chromomere pairs near the centromere,
one prominent pair in the distal one-third region followed
by one small pair, and two more pairs near the knob. This
arm was more heterochromatic than the long arm in chromo­
some 8-D. The short arm was 8.0 um, with two prominent
chromomere pairs near the centromere and one additional
prominent pair slightly distal. This arm was less
heterochromatic than the short arm in chromosome 8-D.
Chromosome 9-D:
The average length of the
chrom o s o m e w a s 2 2 . 2 u m a n d r e l a t i v e l e n g t h w a s 2 . 3 8 .
The centromere was submedian, with an arm ratio of 1.50.
The short arm had a terminal knob and the long arm usually
had nothing or a small chromomere on the end. In FB^9 and
R35P6 the long arm had a small knob or prominent
chromomere. The long arm was 13«3 um, with two prominent
chromomere pairs near the centromere and four small pairs
39
further up the arm. The short arm was 8.9 um, with three
prominent chromomere pairs near the centromere.
Chromosome 9-D' s
This chromosome had an average
length of 22.5 um and a relative length of 2.41. The
centromere was submedian, with an arm ratio of 1.53« Both
arms had terminal knobs. The long arm was I3.6 um, with
two prominent chromomere pairs next to the centromere and
four more pairs distal to these. This chromosome was much
more heterochromatic than chromosome 9-D.
Figures 12A and B represent spreads from Tifgreen,
a commercial hybrid triploid (2n=27) bermudagrass from the
cross C. transvaalensis (2n=18) x C. dactylon var. dactylon
(2n=36). In both spreads there were univalents and
bivalents. In the bivalents there were areas in which the
chromomere patterns did not match, which may or may not be
an artifact of preparation. However in 12A one short arm of
chromosome 3 appeared to be shorter than the other and in
12B on the unpaired terminals of chromosome 2 one
appeared knobbed and the other knobless.
Figures 13 and lb show accessory chromosomes often
found in C. dactylon var. dactylon. In Figure 13 at least
eight B chromosomes could be identified, including a B
trivalent. These B chromosomes were typical of their
appearance in most pachytene spreads, heterochromatincentromere-heterochromatin. Figure 14 shows one typical
Figure 12. Pachytene spreads from Tifgreen showing
bivalents and univalents.—A. One short arm of chromosome
3 appears shorter than the other. B. On the unpaired
terminals of chromosome 2 one appears knobbed and the
other knobless.
Figure 1 3 . Accessory chromosomes from T 3 2 with eight B
chromosomes, including a B trivalent.
Figure 14. Accessory chromosomes from T51 with a B
chromosome, two telo B chromosomes and a fragment.
40
A
101J
8
Figure 12. Pachytene spreads from Tifgreen showing
bivalents and univalents.
B trivalent
Figure 13. Accessor y
chromosomes from TJ2
Figure 14. Accessory
chromosomes from T51
B chromosome pair, two telo or iso B chromosomes and
another fragment of a chromosome without the typical
appearance of a B chromosome.
Somatic Chromosome Morphology
In the diploids, C. dactylon vars. aridus and
afghanicus (2n=18), each chromosome could be distinguished
on the basis of size, arm ratios, and heterochromatic
regions if the spread was in prophase or prometaphase.
In metaphase the only clearly distinguishable chromosome
was the NOR chromosome. Due to its long secondary
constriction, at this stage it was usually the longest
chromosome. The chromosomes could also be matched with
their corresponding pachytene bivalent, as described by
Brilman et al. (1981), based on the appearance of
distinguishing heterochromatic regions.
In Figure 15A the chromosomes are shown, with an
idiogram below showing the characteristic position of
heterochromatin for each chromosome. The number of the
corresponding pachytene bivalent was used to designate
each chromosome. Table 3 shows the chromosome lengths used
to draw the idiogram with the number of the chromosome as
shown. The averages are from 2k chromosomes for the
diploid and 20 chromosomes for the tetraploid.
In the tetraploid, C. dactylon var. dactylon
(2n=36), at prophase each chromosome could be distinguished
42
A
1
2
3
6
5
20
20'
30
7
4
9
8
60
so·
50
8
10
==
DCJ
=
so'
70
70'
40
=;=
40'
90
go'
=
D
80
so'
~
SJJ
Figure 15. Somatic chromosomes with an idiogram below.-A. Diploid var. aridus. B. Tetraploid var. dactylon.
Table J.
Mean values and ranges for somatic chromosome lengths and arm ratios
from diploid (d) and tetraploid (t) clones.
Length (um)
Chromosome
Number
1
Ploidy
Level
Whole
Chromosome
Short arm
Long arm
Arm ratio
Relative
Length
2.7-0.9
(1.5-4.3)
1.0*0.4
(0.4-1.6)
1.7-0.5
(0.9-2.7)
1.85-0.43
(1.23-3.00)
(1.00-1.13)
2.3*1.4
(1.2-4.4)
(0.4-2.0)
0.9*0.7
1.3*0.7
(0.7-2.4)
(1.10-2.25)
d
3.0*1.2
(1.5-6.1)
1.1*0.4
(0.5-1.9)
2.0*0.9
(1.0-4.4)
1.88*0.50
1.15*0.14
(1.11-2.88) (1.00-1.42)
t
2.7*1.6
(1.4-5.2)
1.0*0.7
(0.4-2.0)
1.7*0.9
(0.9-3.2)
1.80*0.39
1.27*0.19
(1.40-2.50) (1.08-1.58)
d
3-3*1.3
(1.5-6.3)
1.1*0.4
(0.5-2.0)
2.1*0.9
(0.9-4.5)
1.96*0.43
(1.07-2.63)
(1.00-1.57)
2.9*1.7
(1.5-5.3)
(0.5-2.0)
1.8*1.0
(1.0-3.3)
1.76*0.19
(1.50-2.00)
(1.15-1.67)
3.6*1.3
1.4*0.6
(0.6-3.1)
2.2*0.8
(1.1-3.9)
1.72*0.48
1.35-0.15
(1.8-6.4)
(1.08-2.83)
(1.10-1.69)
3.2*2.1
(1.5-6.4)
1.1*0.8
(0.4-2.4)
(0.9-4.3)
d
t
2
3
t
6
d
t
1.1*0.6
2.0*1.3
1.55*0.30
1.02±0.04
1.04*0.04
(1.00-1.10)
1.22*0.15
1.36*0.20
1.86*0.41
1.44*0.24
(1.33-2.75) (1.15-1.75)
-p-
Table 3 .
Continued
Length (urn)
Chromosome
Number
5
7
4
9
8
Ploidy
Level
Relative
Length
Whole
Chromosome
Short arm
Long arm
d
3.8*1.4
(1.9-6.5)
1.3-0.7
(0.6-3.2)
2. 6 -0. 9
(1.2-4.5)
2.21*0.71
1.46*0.23
(1.06-3.45) ( I . I O - I . 9 6 )
t
3.3-2.0
(1.6-6.1)
1.2*0.7
(0.6-2.5)
2.1*1.3
(1.0-4.0)
1.70*0.27
1.51*0.23
(1.33-2.22) ( 1 . 2 3 - 1 . 8 3 )
d
4.4*1.7
(1.9-7.9)
1.7*0.7
(0.7-3.1)
2.7±1.0
(1.2-4.8)
1.62*0.42
1.64*0.25
(1.03-2.75) (1.25-2.00)
t
3.7^.5
(1.7-7.4)
1.5-0.9
(0.7-3.2)
2. 2-1. 6
(1.0-4.5)
1.48*0.22
(1.25-2.00)
d
4.4*1.2
(2.8-6.8)
1.7-0.6
(0.8-3.2)
2.7-0.7
(1.7-4.0)
1.61*0.39
1.69*0.23
(1.11-2.88) (1.28-2.14)
t
4.0-2.0
(2.2-7.7)
1.3±0.8
(0.5-2.8)
2.6-1.5
(1.2-5.5)
2.01*0.95
2.03*0.43
(1.19-4.00) (1.30-2.75)
d
5.0*2.0
(2.3-9.3)
1.9*0.9
(0.8-3.8)
3.0*1.2
(1.5-6.0)
1.70*0.50
(1.11-3.00)
Arm ratio
1.66*0.24
(1.31-1.92)
1.87*0.21
(1.55-2.19)
t
2.1*1.7
4.8-3.7
(1.9-H.O) (0.8-5.4)
2.8*2.2
(1.1-6.8)
2.06*0.43
1.39*0.24
(1.07-1.80) (1.54-2.75)
d
5.4-2.1
(2.6-9.7)
2.1*0.9
(0.8-4.3)
3.3*1.3
(1.7-5.7)
1.59*0.28
2.05*0.26
(1.20-2.25) (1.65-2.49)
t
4.4-2.6
(2.0-8.5)
1.8-1.1
2.6*1.6
(1.1-4.8)
1.46*0.22
2.07*0.41
(1.18-1.82) (1.54-2.67)
(0.9-3.9)
ij-5
on the basis of size, arm ratios, and heterochromatic
regions and matched with its corresponding pachytene
bivalent as described in Pachytene Morphology in Tetraploid
C. dact.ylon (Figure 15B). However, by prometaphase the
chromosome number could be determined but not the genome.
Therefore, Table 3 shows the chromosome lengths for each
chromosome number, but not for each genome. However, even
at metaphase chromosome 4-D', the NOR chromosome, appeared
to have a smaller satellite than in the diploid chromosome
k or tetraploid chromosome U~D.
Production of Polyhaploids
Anther culture was not successful in producing
callus and/or polyhaploid plantlets on any of the media
tested. Jeff Krans of Mississippi State University
(personal communication) has produced callus from
vegetative sections of bermudagrass on an MS medium, but
has as yet been unable to achieve regeneration. A MS medium
supplemented with 2,4-D instead of IAA may be more
successful in anther culture of this grass.
Screening of seedlings showed approximately one
twin per thousand seedlings. Out of six sets of twins only
one plant was definitely polyhaploid and this plant
chimeric for tetraploid and for polyhaploid tissue
from the first time the root tips were examined. At
each successive examination more of the cells were
46
tetraploid. A polyhaploid and a tetraploid somatic spread
are shown in Figure 16.
Analysis of meiotic figures was more difficult as
this plant did not flower until more than a year after it
was first selected. Figure 17A shows a diplotene-diakinesis
spread found at this time that appears to represent the
polyhaploid state. Two normal bivalents were observed. The
NOR chromosome formed a bivalent with a loop present
showing an unpaired region. This may be indicative of the
smaller satellite seen in somatic chromosomes for
chromosome 4-D*. Four univalents were present. The other
bivalents showed paired and unpaired regions. Figure 17B
shows a tetraploid diplotene-diakinesis spread found in
another flower. It was difficult to analyze pachytene at
either ploidy level because the chromosomes were sticky.
Figure 18 shows a pachytene with many univalents and
unpaired regions.
Giemsa C-banding
The only treatment that resulted in banding on
the chromosomes was a BSG treatment using barium
hydroxide at room temperature for 50 minutes and 2xSSC
at 60°C for 30 minutes. This resulted in the knobs of the
chromosomes staining darkly with light staining in other
areas with heterochromatin (Figure 19A and B). The bands
revealed little additional information on the
47
A
Figure 16. Somatic spreads from T21.--A.
B. Tetraploid.
Polyhaploid.
u
u
u
A
b
-
b
B
Figure 17. Diplotene-diakinesis spreads fro m T2 1 .-A. Polyhaploid. B. Tetraploid.
48
Figure 18. Pachytene spread from T21 showing univalents
and bi valents.
10JJ
A
B
Figure 19. Giemsa C-bands from Yakima.--A.
B. Pachytene.
Diakinesis.
heterochromatic regions identified by using standard
acetocarmine squash in this species. It did demonstrate
that the prominent chromomeres on the ends of certain
chromosomes are actually small knobs since they stained
the same as the large knobs.
DISCUSSION
Cynodon dact.ylon var. dactylon (2n=36) appears to
be a segmental allotetraploid, with two closely related
genomes, D and D'. In all clones in this study, from
divergent sources, two sets of pachytene bivalents were
present that could be separated by characteristic
morphological features. This supports the suggestion of
segmental allopolyploidy based on metaphase I pairing in
the tetraploid and triploid hybrids, var. dact.ylon
(2n=36) x var. aridus (2n=l8), by Forbes and Burton ( 1 9 6 3 ) ,
Sengupta (1968), and Tripathi et al. (1977).
One set of these chromosomes, D, could be
homologised with the pachytene complement of the diploids
C. dactylon vars. aridus and afghanicus (2n=18) as
described by Brilman et al. ( I 9 8 I ) and Ourecky ( I 9 6 3 ) .
Var. aridus is the only diploid in the genus with
rhizomes and the variety from which other researchers
(Harlan and de Wet 1969; Gupta and Srivastava 1970; Rawal
and Chedda 1971) have suggested var. dactylon arose by
allopolyploidy. Harlan, de Wet, et al. (I970) also
considered var. elegans to be an autotetraploid of var.
aridus.
50
51
In prophase somatic spreads of var. dactylon two
sets of chromosomes could also be separated bycharacteristic morphological features. In both the
diploid, var. aridus, and the tetraploid the somatic
chromosomes could be homologised with their corresponding
pachytene bivalents. The chromosomes were of different
relative lengths during the two phases. The NOR
chromosome was usually the fourth longest at pachytene and
seventh longest in the somatic chromosomes, although in
the diploid it was often the longest due to the amount
of heterochromatin and the long secondary constriction. In
PIantago ovata L. (Hyde 1953)« Lycopersicum
esculentum L. (Rammana and Prakken 1967), and Phaseolus
mungo L. (De and Krishnan 1966) pachytene bivalents and
somatic chromosomes could also be identified and
homologised due to characteristic achromatic and chromatic
parts. Rammana and Prakken (1967) found that chromatic
parts in somatic tissue were contracted by a factor of
4 or 5 over pachytene contraction, whereas achromatic
parts were contracted by a factor of JO. The NOR
chromosome is primarily heterochromatic which would
explain its longer relative length in somatic spreads. The
other chromosomes that changed relative position are
chromosomes 5 and 6 in diploid and tetraploid somatic
spreads and bivalents 8 and 9 in diploid somatic spreads.
52
In each case 5 and 8 have more heterochromatin than
6 or 9 which may explain their lesser contraction.
The pairing behavior in the polyhaploid shows
partial homology between the genomes because of bivalent
formation. The univalents suggest that the homology is not
complete. Polyhaploids from autopolyploids often show
complete synapsis as reported in the polyhaploid from autotetraploid Medicago sativa L. (Gillies and Bingham 1971)
and Sorghum halepense x S. vulgare var. sudanense (Daura
and Stebbins 1952). In autohexaploid Phleum pratense
(2n=6x=42), the polyhaploid makes seven bivalents and seven
univalents at first meitic metaphase, even though each
chromosome is triplicated (Nordenskiold 19^5)- Most
polyhaploids from allopolyploids have a low level of
bivalent formation, which is generally assumed to be due to
the association of corresponding homoeologous chromosomes
from the compoenent genomes (Huskins 1931). If more spreads
at the polyhaploid level were available it may be possible
to determine where between these two extremes the poly­
haploid of var. dactylon lies. The reversion from the
polyhaploid state back to the tetraploid state has also
been reported in Beta vulgaris L. (Kruse 1980) and from
monoploid to diploid in maize (Chase 1 9 5 2 ) .
53
The occasional formation of quadrivalents in the
tetraploid and the overall similarity in the chromosome
measurements of the two sets of pachytene bivalents
suggests the genomes are homoeologous. The origin of the
second set is uncertain. Rawal and Harlan (1971) suggest
a Pleistocene evolution of var. dactylon in India and the
Near East. Earlier a diploid form evolved rhizomes and
invaded the more arid regions of Africa, the Near East,
and India and var. aridus is the modern survivor. Sengupta
(1968) suggested that 0. transvaalensis (2n=18), C.
incompletus (2n=18, rarely 36), C. aethiopicus (2n=18,36),
and C. nlemfuensis (2n=18,36) are allied to C. dactylon
vars. aridus and afghanicus on the basis of pairing in
metaphase I in hybrids, with DD genome. C. transvaalensis
may be D'D' on the basis of triploid pairing behavior in
the cross x var. dactylon. The triploid set would be D'D'D
with some trivalents due to partial homology. The nine
trivalents reported by Forbes and Burton (19^3) would be
due to chance heterogenomic pairing. Harlan, de Wet ,
et al.(l970) found C. incompletus and C. aethiopicus to
be effectively separated from other taxa and 0.
nlemfuensis var. nlemfuensis (2n=18, rarely 36) to be
closely related to C. dactylon vars. aridus and
afghanicus. 0. nlemfuensis and var. aridus may interact
genetically in Southern Ethiopia. C. transvaalensis
could be considered a variet of C. dactylon except it is
morphologically distinct and shows no introgression with
sympatric taxa.
Three possibilities exist for the origin of the
second set of pachytene bivalents:
1) A closely related
diploid species crossed with var. aridus to form a hybrid
after which chromosome doubling occurred. Probable
diploids are C. transvaalensis, with triploid pairing as
above, C. nlemfuensis var. nlemfuensis. C. incompletus,
C. aethiopicus or an extinct diploid. All of these
diploids are native to Africa (Rawal and Harlan 1971)
which would suggest an African origin of C. dactylon var.
dactylon or that one diploid had a different range in the
Pleistocene or late Tertiary. 2) Two distinct cytotypes of
var. aridus or var. afghanicus, not yet identified,
crossed followed by chromosome doubling and diploidization.
Harlan and de Wet (I969) suggested C. dactylon has
populations with different chromosomal behavior and the
gene pool is fragmented with occasional crossing between
subpopulations. 3) An autopolyploid origin with almost
complete diploidization and possibly a gene controlling
pairing as suggested by Thomas and Murray (1978).
Pachytene analysis of other diploids in the genus needs to
be done to further clarify the possibilities.
55
The pachytene bivalents of the tetraploid are much
shorter than the corresponding bivalents in the diploids,
var. aridus and afghanicus, as reported by Brilman et al.
(1981). The longest bivalents in var. dactylon from R35P6
are shorter than the shortest bivalents in the diploids,
which were entering diplotene. Gillies and Bingham (1971)
found in Medicago sativa that the chromosome lengths of the
diploid are considerably longer than the tetraploid or 2x
haploids. That the 2x haploids had pachytene chromosomes of
lengths similar to the tetraploid suggested to them that
greater chromosome contraction may be a property of the
tetraploid state which is still present in the 2x haploid
state. They believed genetic factors causing contraction
may be on the chromosomes. The contraction in the polyhaploid of C. dactylon was not determined. The clone CI 1
had even shorter chromosomes than the other tetraploids and
perhaps this is due to an additional dose of a genetic
factor. Simple increase in chromatin material does not
appear to explain the greater contraction because T32 with
eight accessory chromosomes did not show greater
contraction.
Giemsa C-banding provided no additional
chromosome differentiation. De Jong and Oud (1979) found
in Beta vulgaris that C-banding revealed that the densely
stained parts consist of constitutive heterochromatin and
56
confirmed the striking correspondence between the pachytene
pattern of chromatin segments and the G-banding pattern of
mitotic metaphase chromosomes. Different classes of
heterochromatin in plants appear to be stained in different
C-banding techniques, one pattern consisting of mainly
intercalary/terminal bands and the other centromeric bands
(Filion and Blakey 1979; Yen and Filion 1977). The type
produced in this study was the terminal type. This
corresponded to the knobs found in acetocarmine squash
supporting the theory that they are constitutive
heterochromatin. Correspondence between pachytene and
mitotic heterochromatin was shown using standard
acetocarmine squash in this study.
SUMMARY
Cynodon dactylon var. dactylon appears to be a
segmental allotetraploid. The pachytene bivalents could
all be identified by knobs, prominent chromomeres, lengths
of the chromosomes, long arms and short arms, arm ratios
and relative lengths. These bivalents could be separated
into two sets, D and D', and D could be homologised with
the pachytene bivalents in the diploids C. dactylon vars.
aridus and afghanicus.
In prophase somatic spreads two sets of
chromosomes could be observed in the tetraploid. In
diploid and tetraploid prophase somatic spreads the
chromosomes could be homologised with their corresponding
pachytene bivalents. The chromosomes were of different
relative lengths during the two phases.
The appearance of univalents and bivalents in a
diplotene- diakinesis spread of a polyhaploid of var.
dactylon supports a segmental allopolyploid origin. The
polyhaploid reconverted to a tetraploid before further
studies could be made.
Giemsa C-banding produced telomeric bands in the
position of the knobs seen in standard acetocarmine
squash. The technique that produced the bands was barium
58
hydroxide at room temperature for 50 minutes and 2xSSC at
60°C for 30 minutes to pretreat the slides before Giemsa
staining.
APPENDIX 1
PACHYTENE CHROMOSOME MEASUREMENTS
FROM NINE TETRAPLOID CLONES OF
CYNODON DACTYLON YAR. DACTYLON
59
Table 1.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clonesi
Chromosome 1-D.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
9.1-1.0
(8.3-10.8)
3.8^0.6
(3.1-^.7)
5.3-0.8
(4.2-6.1)
1.04-0.04
1.41^0.36
(1.02-1.97) (1.00-1.10)
CL1
(2)
6.0^1.3
(5.0-6.9)
2.4-0.8
(1.8-2.9)
3.6^0.6
(3.2-4.0)
1.58-0.28
(1.38-1.78)
FBIil-1
(2)
9.9-1.5
(8.8-10.9)
4.2^0.5
(3.8-4.5)
5.7-1.0
(5.0-6.4)
1.13-0.16
1.37-0.07
(1.32-1.42) (1.01-1.24)
9946a
(7)
7.8-0.9
(6.0-8.9)
3.1-0.2
(2.9-3.6)
4.6^0.7
(3.1-5.3)
1.49-0.20
1.01^0.03
(1.07-1.65) ( 1 . 0 0 - 1 . 0 7 )
FB49
(12)
9.7-1.3
(8.0-11.9)
4.2^0.7
(3.1-5.1)
5.5-0.7
(4.7-7.1)
1.32±0.17
1 . 0 3 ^ 0 .04
(I.08-I.58) (1.00-1.11)
R35P6
(6)
12.6^1.0
(11.1-14.0)
5.3-0.7
(4.3-6.3)
7.3-1.1
(6.0-9.0)
1 .04^ 0 . 0 5
1.43^0.36
(I. 0 3 - 2 . 0 9 ) (1.00-1.11)
R7P5
(4)
9.5-1.1
(8.6-11.0)
4.1^0.4
(3.6-4.6)
5.5-0.8
(5.2-6.4)
1.18-0.11
1.35-0.14
(1.15-1.44) (1.08-1.31)
R2P5
(6)
10.6^1.0
(9.4-11.9)
4.0^0.2
(3.7-4.2)
6.6-0.9
(5.7-8.0)
1.68^0.23
1.04-0.03
( I . 3 8 - 2 . 0 5 ) (1.00-1.08)
10.6^2.0
(8.9-13.6)
4.2^1.2
(2.7-5.4)
6.4^1.1
(5.3-8.5)
( 1 . 1 3 - 2 . 3 0 ) (1.00-1.11)
9.8^1.9
4.2^0.9
5.8-1.2
R17P4
Average
(6)
(50)
Arm ratio
1.63^0.46
1.46^0.29
1.00
1.02-0.04
1.04^0.06
Table 2.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromsome 1-D'.
Length (um)
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
9.3-1.3
(8.0-11.4)
3.4^0.5
(3.0-4.3)
5.9-0.9
(4.6-7.1)
1.06±0.08
1.72-0.24
(1.35-2.00) ( 1 . 0 0 - 1 . 1 7 )
CLl
(2)
6.3-1.8
(5.0-7.5)
2.4^0.6
(1.9-2.8)
3.9-1.1
(3.1-4.7)
1.66-0.04
(I.63-I.68)
FB141
(2)
10.4-0.8
(9.8-10.9)
4.3-0.3
(4.1-4.5)
6 .I- 0 . 5
(5.7-6.4)
1.41-0.02
1.19-0.08
(1.39-1.42) (1.13-1.24)
9946a
(7)
7.9-1.1
(6.1-9.2)
3.2^0.4
(2.8-3.8)
4.7-0.8
(3.2-5.4)
1.02-0.04
1.47-0.19
(1.10-1.67) (1.00-1.12)
FB49
(12)
9.8-1.4
(8.0-12.2)
4.2^0.8
(3.0-5.4)
5.6-0.9
(4.2-7.1)
1.04±0.04
1.36-0.26
(1.00-1.77) (1.00-1.10)
12.6-0.4
(II. 9 - I 3 .O)
4.7-0.5
(3.9-5.2)
(7.6-8.0)
1.67-0.21
1.04^0.05
(1.50-2.05) (1.00-1.11)
8.4-0.6
(8.0-9.2)
3 . 6^0.5
(3.0-4.1)
4.8^0.6
(4.3-5.6)
1.04^0.08
1.36^0.30
(1.05-1.67) ( 1 . 0 0 - 1 . 1 5 )
10.2-0.8
4.1-0.1
(3.9-4.2)
6.2^0.7
(5.0-6.8)
1.0l±0.01
1.53-0.15
(1.28-1.63), (1.00-1.02)
1.02^0.03
1.71-0.72
(
1
.00-1.07)
(1.07-3.09)
R35P6
(6)
R7P5
(4)
R2P5
(6)
(8.9-H.O)
R17P4
Average
(6)
(50)
7.8-0.2
10.6^1.8
(8.9-13.1)
(3.2-5.8)
6.5-1.8
(5.0-9.9)
9.7-1.9
3.9-0.8
5.8^1.3
4.0^0.9
Arm ratio
1.53-0.33
Relative
Length
1.05-0.06
(1.00-1.09)
1.04^0.06
Table J.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 2-D.
Length (um)
Clone
No.
Cells
B296
(5)
10.8^1.4
(9.4-13.0)
4.1^0.8
(3.0-5.0)
6.8^0.8
(5.9-8.0)
1.23^0.04
1.69^0.25
(1.52-2.13) (1.18-1.27)
Cll
(2)
7.3-1.1
(6.5-8.0)
3.1^1.1
(2.3-3.8)
4.2
1.23^0.10
1.47^0.52
(1.10-1.83) (1.16-1.30)
FB141
(2)
12.0-2.8
(10.0-14.0)
4.6^0.7
(4.1-5.1)
7.4±2.1
(5.9-8.9)
1.60^0.22
1.37-0.31
(1.44-1.75) (1.15-1.59)
9946a
(7)
9.4±1.6
(7.9-11.8)
4.1^0.5
(3.4-4.8)
5.3-1.1
(4.0-7.0)
1.29-0.19
1.23^0.15
(1.03-1.46) (1.04-1.44)
FB49
(12)
11.6^1.5
(9.7-14.1)
4.2^0.6
(3.3-5.5)
7.3-1.3
(5.9-9.9)
1.24-0.13
1.75-0.38
(1.20-2.36) (1.03-1.44)
R35P6
(6)
13.7-1.3
(12.8-16.0)
5.4^0.7
(4.3-6.0)
8.4^1.3
(7.4-10.2)
1.6l±0.42
1.13^0.09
(1.13-2.28) (1.03-1.27)
R7P5
CO
11.oil.2
(9.3-11.9)
3-9^1.0
(3.1-5.1)
7.1^1.0
(6.1-8.5)
1.90^0.61
1.36^0.14
(1.33-2.74) (1.16-1.49)
R2P5
(6)
11.3^0.7
(10.1-12.2)
4.5^0.3
(4.1-4.8)
6.8^0.7
(5.7-7.5)
1.52±0.19
(1.30-1.83)
R17P4
(6)
11.7^2.4
(9.4-15.9)
4.6^0.9
(3.3-5.7)
7.1^1.9
(6.1-10.8)
1.13±0.09
1.58^0.41
(1.11-2.12) (1.06-1.29)
(50)
11.2^2.0
4.4^0.8
6.9-1.5
Average
Whole
Chromsome
Short arm
Long arm
Arm ratio
1.61^0.37
Relative
Length
1.11^0.08
(1.01-1.24)
1.21±0.l4
Table 4.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 2-D'.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
10.8±2.3
(8.6-14.4)
4.2±1.0
(2.8-5.4)
6.6^1.4
(5.4-9.0)
1.60-0.29
1.23-0.12
(1.37-2.07) (1.08-1.41)
CLl
(2)
7.3-0.7
(6.8-7.8)
2.8-0.3
(2.6-3.0)
4.5^0.4
(4.2-4.8)
1.6l±0.01
1.25-0.16
(1.60-1.62) (1.13-1.36)
FBI4-1
(2)
10.9-0.1
4.0-0.3
(10.8-11.0) (3.8-4.2)
6.9-0.4
(6.6-7.2)
1.25±0.01
1.73-0.23
(1.57-1.89) (1;24-1.25)
9946a
(?)
3.6-0.5
9.3-1.1
(8.1-11.3) (3.1-4.1)
5.7-0.8
(5.0-7.3)
1.6l±0.23
1.21-0.11
(1.22-1.94) (1.13-1.38)
(12)
11.4-1.2
4.5-0.7
(10.0-14.1) (3.6-5.8)
6.9±0.8
(5.9-8.3)
1.23^0.16
1.57-0.28
(1.27-2.31) (1.03-1.48)
(6)
14.5^1.4
5.2-0.8
(12.7-16.1) (4.8-5.9)
9.3-1.3
(8.8-11.3)
1.82±0.4l
1.20^0.14
(1.32-2.35) (1.05-1.43)
R7P5
CO
10.9-1.4
4.2-0.7
(9.4-12.2) (3.5-5.3)
6.7-1.3
(5.5-8.4)
1.62^0.52
1.35-0.16
(1.30-2.40) (1.18-1.53)
R2P5
(6)
11.5^0.6
4.9-0.6
(11.0-12.1) (4.0-5.6)
6.7-0.4
(6.1-7.1)
1.40±0.24
1.14±0.0 6
(1.14-1.75) (1.09-1.24)
R17P4
(6)
5.0±2.0
12.0^2.6
(9.7-16.3) (2.9-8.1)
7.1-1.1
(5.2-8.2)
1.60±0.60
1.16^0.09
(1.01-2.34) (1.09-1.33)
FB49
R35P6
Average
(50)
11.3-2.1
4.4^1.1
6.9-1.4
Arm ratio
1. 6 1 ^ 0 . 3 5
1.21±0.13
Table 5»
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 3-D.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
11.1-2.9
(7.7-14.8)
3-2^0.9
(2.1-4.1)
8.0-2.0
(5.6-10.8)
1.26-0.23
2.56-0.24
(2.17-2.75) (1.00-1.61)
CLl
(2)
7.3-0.2
(7.1-7.4)
2.3-0.6
(1.9-2.7)
5.0-0.4
(4.7-5.2)
2.24-0.71
1.26-0.32
(1.74-2.74) (1.03-1.48)
FBI41
(2)
9.8-1.4
(8.8-10.8)
2.6-0.1
(2.5-2.6)
7.3-1.3
(6.3-8.2)
2.84-0.45
1.12-0.17
(2.52-3.15) (1.00-1.24)
99^6a
(7)
10.0-2.3
(7.8-14.0)
2.5-0.5
(1.8-3.3)
7.5-1.8
(5.6-10.7)
3.08^0.38
1.30^0.25
(2.33-3.38) (1.01-1.77)
FB49
(12)
11.8-0.8
(10.9-13.6)
2.9-0.4
(2.5-3-8)
8.8-0.9
(7.2-10.5)
04-0.49
1.27-0.15
('-1.05-3.84) (1.04-1.51)
R35P6
(6)
15.0^1.8
(13.4-18.1)
3.1-0.4
(2.9-3.9)
11.9-1.4
(10.5-14.2)
1.24^0.15
3.79-0.28
(3.57-4.30) (1.09-1.43)
R7P5
(4)
11.1^2.2
(8.0-13.0)
2.9-1.0
(1.9-4.3)
8.2±1.4
(6.1-9.0)
3.02-0.72
1.37-0.26
(2.02-3.75) ( 1 . 0 0 - 1 . 6 3 )
R2P5
(6)
12.8-1.7
(10.8-14.7)
3.0^0.7
(2.2-3.9)
9-8-1.5
(8.0-11.7)
1.27-0.20
3.38^0.83
(
I.O3-I.63)
(2.05-3.96)
11.7-1.3
(9.4-13.2)
3.1-0.4
(2.6-3.6)
(6.8-9.9)
11.7-2.3
2.9-0.6
8.7-2.0
R17P4
Average
(6)
(50)
8.6-1.1
Arm ratio
1.14^0.12
2.81-0.29
(2.36-3.10) ( 1 . 0 0 - 1 . 3 3 )
3.06-0.59
1.26-0.19
Table 6.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 3-D'.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
11.0-2.2
(8.0-12.9)
3.9-1.2
(2.2-5.3)
7.1-1.2
(5.8-8.5)
1.24^0,15
1.95-0.44
(1.42-2.64) (1.04-1.40)
CLl
(2)
7.2
2.4^0.6
(2.0-2.8)
4.8^0.6
(4.4-5.2)
1.24±0.28
2.09-0.73
(1.04-1.44)
(1.57-2.60)
FBI41
(2)
9.9-0.4
(9.6-10.1)
3.0^0.6
(2.5-3.4)
6.9-1.0
(6.2-7.6)
2.43^0.86
1.13^0.05
(1.82-3.04) (1.09-1.16)
9946a
(7)
9.7-1.8
(8.0-12.0)
3.1-0.5
(2.5-4.0)
6.5-1.7
(4.8-8.7)
2.11±0.59
(I.50-2.90)
1.26-0.16
(1.03-1.52)
FB49
(12)
12.5±1.3
(10.0-14.5)
3.8±0.6
(3.0-4.6)
8.8-1.3
(6.3-10.7)
2.40^0.61
(1.67-3.57)
(1.12-1.65)
Arm ratio
1.34^0.16
R35P6
(6)
15.0±1.6
(12.6-17.2)
4.5-1.0
(3.3-4.8)
10.5-1.6
(9.3-13.6)
1.24-0.17
2.47-0.77
(1.63-3-78) (1.04-1.55)
R7P5
(4)
11.1-2.2
(8.0-13.0)
3.5-1.0
(2.1-4.3)
7.6^1.2
(5.9-8.7)
1.37-0.27
2.29-0.37
(2.02-2.81) (I.00-I. 6 3 )
R2P5
(6)
12.8^1.4
(11.0-15.2)
3.8^0.7
(2.7-4.7)
9.0-1.5
(6.8-11.2)
1.26^0.13
2.47-0.71
(1.62-3.37) (1.13-1.45)
12.8^2.0
(10.7-16.4)
4.6^1.0
(3.1-5.6)
8.2^2.5
(6.0-12.9)
1.23^0.09
1.94^0.97
(1.10-1.33)
(1.15-3.69)
3.8-0.9
8.1^2.5
R17P4
Average
(6)
(50)
11.9-2.4
2.26^0.65
1.27-0.16
Table 7.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 4-D.
Length (urn)
Clone
No.
Cells
Whole
Chromosome
B296
(5)
10.7-0.9
Arm ratio
Relative
Length
Short arm
Long arm
(9.8-11.7)
4.4^0.9
(3.3-5.5)
6.3-0.5
(5.6-6.8)
1.24-0.23
1.^0.35
(1.13-2.06) (1.00-1.52)
CL1 (*)
(2)
7.7-0.8
(7.1-8.3)
3.5-0.6
(3.0-3.9)
^.3-0.2
(4.1-4.4)
1.31^0.16
1.25-0.17
(1.13-1.37) (1.20-1.42)
FB141 (*)
(2)
9.9-1.6
(8.7-H.O)
^.5-0.6
(4.0-4.9)
5.4±1.0
(4.7-6.1)
1.21±0.40
1.13^0.18
(1.18-1.24) (1.00-1.25)
9946a (*)
(7)
11.7-1.3
(10.2-14.0)
4.6^*0.5
(4.0-5.5)
7.1-1.1
(5.9-8.5)
1.56^0.27
1.55^0.22
(1.30-2.13) (1.15-1.80)
(11)
11.2^1.6
(9.6-13.9)
^.5-0.9
(3.5-6.1)
6.7-1.0
(5.8-8.6)
1.51^0.26
1.23^0.19
(1.11-2.02) (1.00-1.74)
(6)
11.6-2.4
(9.1-16.0)
4.6^0.9
(3.0-5.5)
7.1-2.0
(5.5-10.9)
0.96-0.17
1.59-0.47
(1.12-2.14) (0.80-1.27)
R2P5
(5)
13.0^1.9
(9.7-1^.5)
5.7-0.8
(4.4-6.6)
7.3-1.4
(5.3-9.0)
1.30^0.22
1.29-0.21
(1.05-1.64) (1.00-1.52)
R17P4 (*)
(6)
14.1^2.4
(11.0-17.1)
6.2^1.2
(5.0-8.0)
7.9-1.5
(6.0-9.4)
1.30^0.29
1.37-0.13
(1.13-1.88) (1.24-1.60)
11.9-2.7
5.0-1.3
6.9-1.7
FB49
R35P6
R7P5
Average (*) (17)
1.39-0.28
1.31-0.22
Table 8.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 4-D'.
Lenght (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
13.3-1.6
(11.8-15.0)
5.6*1.0
(4.2-7.0)
7.7*1.1
(6.0-9.1)
1.52*0.11
1.41*0.33
(1.03-1.81) (1.45-1.70)
CLl
(2)
7.7*0.7
(7.2-8.2)
3.5*0.8
(2.9-4.0)
4.3*0.1
(4.2-4.3)
1.32*0.18
1.27*0.30
(1.05-1.48) (1.19-1.44)
FBI41
(2)
10.5*2.2
(8.9-12.0)
4.9*1.3
(4.0-5.8)
5.6*0.9
(4.9-6.2)
1.15*0.11
1.19*0.24
(1.07-1.23) (1.02-1.36)
99^6a
(7)
12.1*0.8
(11.2-13.4)
5.2*0.5
(4.5-5.9)
7.0*0.8
(5.7-8.2)
1.60*0.21
1.36*0.25
(1.00-1.82) (1.28-1.97)
FB49
(12)
11.8*1.1
(10.2-13.5)
4.9*0.6
(4.3-6.0)
7.0*0.9
(5.8-8.4)
1.43*0.22
1.27*0.19
(1.05-1.75) (1.02-1.65)
R35P6
(6)
17.1*3.3
(13.4-21.9)
7.0*1.5
(5.7-9.8)
10.2*2.3
(7.5-13.5)
R7P5
(4)
11.4*0.5
(11.0-12.1)
4.6*0.6
(3-8-5.1)
6.8*0.6
(6.2-7.5)
1.41*0.03
1.49*0.32
(1.29-1.97) (1.38-1.44)
R2P5
(6)
12.4*1.2
(10.7-13.2)
5.4*0.7
(4.4-6.2)
7.0*0.8
(5.7-8.1)
1.22*0.14
1.31*0.19
(1.13-1.59) (1.10-1.48)
R17P4
(6)
14.2*2.5
(11.0-17.0)
6.0*1.2
(4.6-7.8)
8.2*1.4
(6.2-9.2)
1.38*0.13
1.37*0.14
(1.18-1.53) (1.24-1.63)
5.4*1.1
7.4*1.7
Average
(50)
12.7*2.5
Arm ratio
1.48*0.31
(I. 2 3 - 2 . 0 8 )
1.39*0.24
1,41*0.25
(1.03-1.71)
1.37*0.21
Table 9.
Mean values and. ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 5-D.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
11.8-2.0
(10.0-14.9)
4.0-1.1
(2.9-5.3)
7.8-1.9
(6.8-11.1)
1.34*0.08
2.05*0.71
(1.28-2.92) (1.25-1.46)
CLl
(2)
7.6-0.9
(6.9-8.2)
3.6*0.3
(3.4-3.8)
5.0*0.6
(4.5-5.4)
1.46*0.18
1.37-0.07
(1.32-1.42) (1.33-1.58)
FBI41
(2)
1 3 . 1-1. 5
(12.0-14.1)
5.7-1.9
(4.3-7.0)
7.4*0.4
(7.1-7.7)
1.49*0.16
1.40*0.55
(1.01-1.79) (1.38-1.60)
9946a
(7)
1 0 . l-l.5
(8.1-12.0)
3.7-0.3
(3.4-4.4)
6.4-1.3
(4.7-8.4)
FB49
(12)
13.1-1.8
(10.3-15.5)
4.9-0.8
(3.1-6.0)
8.1*1.9
(5.5-12.3)
1 .74* 0 . 7 6
1.39*0.15
(1.15-3.97) (1.14-1.58)
(6)
16.6-1.6
(15.1-18.8)
6.0*1.2
(4.6-7.8)
10.6*0.5
(9.8-11.1)
1.83*0.34
1.37*0.13
(1.41-2.28) (1.19-1.53)
(10.2-13.6)
4.6-0.8
(3.8-5.6)
7.7-1.6
(5.6-9.3)
1.52*0.17
1.73-0.53
(1.22-2.21) (1.28-1.70)
12.7-0.9
(11.9-14.1)
5.2*1.0
(4.1-6.6)
7.5*0.2
(7.3-7.8)
1.26*0.14
1.50*0.29
(1.12-1.51)
(1.14-1.90)
(10.0-17.0)
5.4-1.1
(3.8-7.0)
8.2-1.8
(6.0-10.0)
1.31*0.11
1.53-0.27
(1.09-1.92) (1.12-1.43)
12.7-2.6
4.8-1.2
7.9-1.9
R35P6
R7P5
(4)
R2P5
(6)
R17P4
(6)
Average
(50)
12.3-1.5
13.6-2.7
Arm ratio
1.72*0.35
(1.38-2.33)
1.72*0.52
1.31*0.13
(1.14-1.47)
1.36*0.14
Table 10.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 5-D'.
Length (urn)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
12.1*2.0
(10.0-15.4)
4.6*1.1
(3.2-5.8)
7.5*1 •8
(6.0-10.2)
1.38*0.10
1.72*0.53
(1.09_2.21) (1.25-1.51)
CLl
(2)
8.3-0.7
(7.8-8.8)
2. 9 *0. 1
(2.8-3.0)
5.4*0.8
(4.8-6.0)
1.42*0.20
1.87*0.38
(1.60-2.14) (1.28-1.56)
FB141
(2)
12.7-2.5
(10.9-14.5)
5.5*2.1
(4.0-7.0)
7.2*0.4
(6.9-7.5)
1.42*0.24
1.40*0.47
(1.07_1.73) (1.25-1.59)
9946a
(7)
10.3*1.1
(8.5-11.8)
4.1-0.8
(3.0-5.1)
6.2*1.0
(5.0-7.7)
1. 6 0 *0. 4 9
1.35*0.13
(1.09-2.40) (1.20-1.44)
FB49
(12)
13.3*1.6
(10.2-15.5)
5.1*1.0
(4.0-7.2)
8.2*1.4
(6.1-11.3)
1.67*0.48
1.42*0.19
(1.10-2.83) (1.13-1.76)
R35P6
(6)
16.6-2.4
(13.7-19.9)
6.0*1.6
(4.4-8.1)
10.6*1.1
(9.3-U.8)
1.84*0.36
1.36*0.19
(1.31-2.13) (1.17-1.58)
R7P5
(4)
11.8-1.6
(9.8-13.7)
3.4-0.9
(2.6-4.3)
8.4*1.0
(7.2-9.4)
1.46*0.20
2.50*0.37
(2.16-2.87) (1.22-1.71)
R2P5
(6)
13.7*1.9
(11.2-16.1)
5.5*0.7
(5.0-6.6)
8.1*1.7
(6.2-10.3)
1.35*0.24
1.49*0.33
(1.05-1.92) (1.12-1.75)
14.1*3.1
(9.9-17.7)
5.8-0.8
(4.7-7.0)
8.3*2.4
(5=2-10.7)
1.40*0.29
1.35*0.15
(1.02-1.72) (1.11-1.51)
12.9*2.7
4.9*1.3
8.0*1.9
R17P4
Average
(6)
(50)
Arm ratio
1.69*0.48
1.39*0.17
Table 11.
Mean values and rnges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 6-D.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
13.5*3.3
(10.0-17.5)
4.4*1.1
(2.9-5-5)
9.1*2.2
(7.1-11.9)
2.12*0.30
1.53*0.22
(1.69-2.45) (1.25-1.78)
CL1
(2)
8.6*0.9
(7.9-9.2)
3.6*0.3
(3.^-3.8)
5.0*0.6
(4.5-5.4)
1.46*0.18
1.37*0.07
(1.32-1.42) (1.33-1.58)
FBI4-1
(2)
13.5*2.3
(11.8-15.1)
5.1*0.4
(4.8-5.3)
8.4*2.0
(7.0-9.8)
1.66*0.28
1.54*0.25
(1.46-1.85) (1.36-1.72)
9946a
(7)
11.5*1.7
(9.0-13.8)
4.0*0.7
(3.1-5.2)
7.4*1.2
(5.8-9.3)
1.51*0.18
1.87*0.33
(1.38-2.27) (1.27-1.75)
FB49
(12)
15.0*1.6
6.1*1.4
(4.3-8.7)
8.9*0.8
(7.5-10.2)
(1.10-2.26) ( I . 3 6 - I . 7 8 )
(13.0-17.9)
Arm ratio
1.52*0.38
1.60*0.15
R35P6
(6)
19.5*2.3
(16.3-21.9)
8.1*1.8
(5.4-10.0)
11.4*1.1
(9.3-12.0)
1.47*0.40
1.61*0.23
(1.16-2.19) (1.25-1.91)
R7P5
(4)
13.5±2.2
(10.8-16,,0)
4.6*1.5
(3.3-6.8)
9.0*1.5
(6.8-10.0)
1.64*0.28
2.10*0.72
(1.35-2.97) (1.25-1.90)
R2P5
(6)
14.4*1.6
(12.5-16.4)
4.9*1.4
(3.2-7.0)
9.5*1.6
(7.2-11.8)
2.12*0.89
1.43*0.22
(1.14-1.75)
(1.33-3.69)
R17P4
(6)
15.2*3.3
(11.8-21.0)
6.6*1.0
(5.3-7.8)
8.6*2.5
( 6 .1- 1 3 . 2 )
1.30*0.26
1.46*0.15
(1.04-1.69) (1.31-1.71)
5.5*1.8
8.9*1.9
Average
(50)
14.4*3.2
1.71*0.54
1.54*0.19
Table 12.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 6-D'.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
13.6*2.8
(10.3-16.7)
4.8*1.1
(3.1-5-9)
8.9*2.0
(7.2-11.7)
1.91*0.41
1.55*0.18
(1.41-2.34) (1.29-1.77)
CLl
(2)
9.1*0.4
(8.8-9.3)
3.8*0.6
(3.3-4.2)
5.3*1.0
(4.6-6.0)
1.46*0.51
1.56*0.29
(1.10-1.82) (1.35-1.76)
FBI41
(2)
13.5*1.6
(12.3-14.6)
5.0*0.l
(4.9-5.0)
8.5*1.7
(7.3-9.7)
1.55*0.16
1.72*0.37
(1.46-1.98) (1.41-1.66)
99^6a
(7)
11.7*1.9
(8.6-14.0)
4.8*0.9
(4.0-6.4)
6.8*1.5
(4.6-9.4)
1. 4 4 *0. 3 9
1.54*0.29
(1.15-2.29) (1.21-1.98)
FB49
(12)
15.4*1.9
(12.1-18.7)
6.6*1.2
(4.4-8.2)
8. 9 *1. 2
(7.2-10.7)
1.65*0.16
1.39*0.30
(1.01-1.91) ( 1 . 3 8 - I . 8 9 )
R35P6
(6)
19.5*2.2
(16.0-22.1)
7.6*1.4
(6.1-9.9)
11.9*2.3
(8.8-15.9)
1.64*0. 5 5
(1.22-2.56)
R7P5
(4)
13.8*2.1
(11.0-16.0)
5.0*0.4
(4.4-5.3)
8.8*2.1
(5.9-10.8)
1.77*0.44
1.70*0.23
(1.16-2.11) (1.38-1.90)
R2P5
(6)
14.7*1.3
(12.9-16.0)
6.5*1.0
(4.9-7.9)
8.2*0.9
(6.6-9.2)
1.29*0.26
1.45*0.20
(0.99-1.63) (1.17-1.76)
R17P4
(6)
16.0*2.9
(13.4-21.0)
6.0*1.3
(4.0-7.7)
10.0*1.8
(8.0-13.3)
1.70*0.35
(1.38-2.35)
1.55*0.10
(1.43_1.71)
14.7*3.1
5.9*1.4
1.54*0.40
1.58*0.20
Average
(50)
8.8*2.2
Arm ratio
1.61*0.20
(1.23-1.81)
Table 13.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 7-D.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
15.9*3.0
(12.6-20.3)
6.7*1.8
(4.6-9.2)
9.2*1.3
(8.0-11.1)
1.41*0.21
1.80*0.17
(1.21-1.74) (1.60-1.99)
CL1
(2)
10.6*0.3
(10.4-10.8)
4.4*0.1
(4.3-4.4)
6.3*0.2
(6.1-6.4)
1.44*0.02
1.83*0.36
(1.42-1.45) (1.57-2.08)
FB141
(2)
14.6*1.6
(13.**-15.7)
6.5-1.7
(5.3-7.7)
8.1*0.1
(8.0-8.1)
1.66*0.17
1.29*0.35
(1.04-1.53) (1.54-1.78)
9946a
(7)
13.3-2.0
(10.6-15.8)
5.4*1.8
(3.0-7.6)
7.9*0.4
(7.4-8.5)
1.62*0.55
1.76*0.39
(1.06-2.53) (1.42-2.47)
FB49
(12)
16.4*1.7
(14.1-19.9)
7.0*1.0
(5.6-9.0)
9.4*1.3
(8.0-12.9)
1.76*0.21
1.38*0.27
(1.01-1.84) ( 1 . 4 3 - 2 . 1 5 )
11.7*1.5
1.81*0.13
1.18*0.35
(0.91-1.87) (1.65-1.99)
R35P6
(6)
22.0-2.0
10.3*1.8
(20.0-25.1) (7.0-12.0)
( 1 0 . 2- 1 3 . 9)
Arm ratio
R7P5
(4)
16.6*2.4
(14.8-20.1)
6.8*1.0
(5.3-7.7)
9.8*1.9
(7.8-12.4)
1.48*0.33
2.05*0.31
(1.11-1.87) (1.85-2.51)
R2P5
(6)
16.4-1.2
(14.8-18.2)
6.3*1.2
(4.6-8.2)
10.1*0.6
(9.1-11.1)
1.62*0.17
1.66*0.35
(1.22-2.22) (1.45-1.91)
R17P4
(6)
18.0*3.8
8.3*2.6
(13.6-22.9) (5.9-11.4)
9.7*1.4
(7.5-11.5)
1.23*0.28
1.73*0.14
(0.91-1.64) (1.53-1.91)
Average
(50)
16.5-3.4
7.1*2.1
9.4*1.7
1.41*0.28
1.77*0.24
Table 14.
Mean values and rnages among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 7-D'.
Length (um)
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
15.3*3-3
(12.1-20.4)
6.4*1.8
(4.9-9.1)
8.9*1.6
(7.2-11.3)
CLl
(2)
10.9*1.3
(10.0-11.8)
4.3*0.6
(3.8-4.7)
6.7*0.6
(6.2-7.1)
Arm ratio
Relative
Length
1.41*0.16
1.73*0.18
(1.24-1.66) (1.57-2.00)
1.57*0.08
(I.5I-I.63)
1.86*0.21
(1.71-2.00)
FB141
(2)
14.9*0.1
(14.8-15.0)
6.2*0.3
(6.0-6.4)
8.7*0.1
(8.6-8.8)
1.41*0.09
1.70*0.03
(1.34-1.47) (1.68-1.72)
9946a
(7)
12.8*1.7
(10.5-15.4)
5.6*1.4
(3.0-7.4)
7.2*0.7
(5.8-8.0)
1.68*0.27
1.39*0.51
(1.04-2.50) (1.43-2.12)
FB49
(12)
17.3*2.2
7.1*1.1
(5.4-8.7)
10.3*1.7
(7.4-13.3)
1.85*0.21
1.49*0.33
(0.95-2.07) (1.52-2.22)
12.9*0.7
(12.1-13.8)
1.50*0.24
1.79*0.13
(1.23-1.76) (1.62-1.97)
(15.2-21.8)
R35P6
(6)
R7P5
W
R2P5
R17P4
Average
(6)
(6)
(50)
8.8*1.3
21.7*1.3
(19.6-23.4) (7.1-10.5)
(15.0-20.1)
7.0*0.7
(5.8-9.2)
10.1*0.8
(9.2-10.9)
2.12*0.28
1.49*0.26
(1.18-1.78) (1.88-2.51)
16.8*1.5
(14.8-19.0)
7.7*0.7
(6.8-8.7)
9.1*0.9
(8.0-10.8)
1.19*0.10
(1.06-1.32)
17.0*3.2
7.5*2.0
(13.1-21.2) (5.0-10.1)
9.5*1.3
(8.1-11.1)
1.64*0.11
1.31*0.21
(1.10-1.62) (1.47-1.72)
17.1*2.1
16.5*3.3
7.0*1.6
9.6*1.3
1.41*0.29
1.66*0.21
(1.47-2.01)
1.78*0.23
Table 15•
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 8-D.
Length (um)
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
18.0*2.5
(16.1-22.0)
7.0*0.8
(6.3-8.0)
11.0*2.1
(9.2-14.0)
CLl
(2)
12.3*0.6
(11.9-12.7)
5.6*0.6
(5.1-6.0)
(6.7-6.8)
6.8*0.1
Arm ratio
Relative
Length
2.05*0.13
1.57*0.28
(1.19-1.92) (1.83-2.16)
1.23*0.15
(1.12-1.33)
2.11*0.38
(1.84-2.38)
FB141
(2)
15.9*1.6
(14.7-17.0)
7.1*0.1
(7.0-7.1)
8.8*1.6
(7.7-9.9)
1.25*0.16
1.81*0.17
(1.10-1.39) (1.69-1.93)
9 946a
(7)
14.6*1.6
(12.1-16.8)
6.4*0.8
(5.1-7.8)
8.2*1.2
(7.0-10.4)
1.28*0.20
1.93*0.30
(1.05-1.63) (1.70-2.50)
(12)
20.0*3.4
8.2*1.7
(16.0-27.6) (5.0-11.6)
11.8*2.1
(8.3-16.0)
2.13*0.26
1.47*0.30
(1.02-2.20) (1.76-2.55)
R35P6
(6)
9.2*1.2
23.7*2.9
(19.8-28.3) (6.9-10.1)
14.5*2.5
(12.1-18.8)
1.61*0.32
1.96*0.27
(1.21-1.98) (1.64-2.25)
R7P5
(4)
19.3*2.2
7.3*2.4
(17.1-22.3) (4.6-10.3)
11.9*0.5
(11.4-12.5)
1.78*0.67
2.39*0.31
(1.17-2.72) (2.04-2.79)
R2P5
(6)
8.0*1.3
19.1*2.5
(15.2-22.1) (6.0-10.0)
11.0*1.6
(9.0-12.6)
1.40*0.23
1.88*0.29
(1.11-1.70) (1.54-2.25)
R17P4
(6)
7.4*1.6
19.7*4.0
(14.8-25.2) (5.8-10.0)
12.3*2.9
(9.0-17.2)
1.68*0.31
1.89*0.13
(1.51-2.15) (1.66-2.05)
FB49
Average
(50)
18.8*3.9
7-6*1.6
11.2*2.7
1.49*0.33
2.02*0.28
Table 16,
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 8-D'.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
17.7*2.9
(15.0-22.4)
7.5*0.5
(6.9-8.2)
10.2*2.4
(7.9-14.2)
2.02*0.23
1.34*0.23
(1.11-1.73) (1.70-2.27)
CLl
(2)
12.4*0.6
(12.0-12.8)
5-5*0.4
(5.2-5.8)
6.9*0.1
(6.8-7.0)
1.26*0.07
2.13*0.38
(1.21-1.31) (1.86-2.40)
FB141
(2)
15.4*1.1
(14.6-16.1)
5.4*0.8
(4.8-6.0)
10.0*0.2
(9.8-10.1)
1.86*0.25
1.76*0.11
(1.68-2.04) (1.68-1.83)
99^6a
(7)
14.8-1.0
(13.1-15.9)
(5.7-8.2)
8.3*1.3
(6.8-9.9)
1.30*0.32
1.96*0.32
(0.83-1.68) (1.70-2.65)
19.9*2.7
8 . 5*1• 3
(16.0-25.1) (6.7-10.7)
11.4*2.3
(8.4-17.1)
2.12*0.38
1.36*0.33
(1.01-2.14) (1.86-2.49)
10.5*2.6
(7.0-13.2)
15.4±2.7
(12.0-20.0)
1.53*0.40
(1.20-2.29)
2.13*0.38
(1.68-2.63)
19.4-3.4
(16.4-24.2)
6.8-0.8
(6.1-8.0)
12.6*2.6
(9.9-16.2)
1.85*0.23
(I.52-2.03)
2.40*0.44
(2.05-3.03)
7.6±1.3
(6.0-9.2)
11.7*1.2
(10.2-13.2)
1.90*0.17
1.57*0.30
(1.29-2.12) (1.65-2.05)
9.5*1.3
(7.8-14.1)
1.64*0.11
1.31*0.21
(1.02-1,67) (1.62-2.19)
FB49
R35P6
(12)
(6)
25.8-4.6
(20.3-33.2)
R7P5
(4)
6.5*0.8
R2P5
(6)
19.3*2.0
(17.1-22.1)
R17P4
(6)
17.0*3.2
7.5*2.0
(14.4-26.9) (6.1-12.8)
Average
(50)
19.2-4.4
8.0*2.0
11.2*1.3
Arm ratio
1. 4 4 *0. 3 4
2.05*0.29
Table 17.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones; Chromosome 9-D.
Length (um)
Whole
Chromosome
Relative
Length
Clone
No.
Cells
B296
(5)
22.8*2.6
9.2*1.7
(19.1-26.0) (8.0-12.0)
CLl
(2)
13.9-1.0
(13.2-14.6)
5.9*1.3
(4.9-6.8)
8.1-0.4
(7.8-8.3)
1.42*0.3 8
(1.15-1.69)
FBI4-1
(2)
16.5-0.1
(16.4-16.5)
5.6*0.4
(5.3-5.8)
10.9*0.3
( 1 0 . 7 -H.I)
1.88*0.03
1.97*0.18
(1.84-2.09) (1.86-1.90)
16.6-2.0
(12.9-18.6)
7.2*0.9
(5.7-8.2)
9.3*1.3
(7. 2 - 1 0 . 9 )
1.29*0.14
2.19*0.34
(1.10-1.54) (1.70-2.68)
Short arm
Long arm
13.7*2.0
(10.2-15.1)
Arm ratio
2.27*0.34
1.53*0.35
(1.14-1.89) (2.17-3.25)
2.42*0.71
(1.91-2.92)
9946a
(7)
FB49
(12)
22.9*4.1
9.1*1.3
(18.0-30.6) (7.7-H.l)
13.8*3.2
(10.3-19.5)
1.52*0.28
2.44*0.3 8
(1.17-2.10) (1.98-3.12)
R35P6
(6)
11.1*1.6
28.2*2.5
(25.1-32.0) (8.5-12.8)
17.1*2.4
(13.9-20.0)
1.58*0.38
2.32*0.17
(1.19-2.09) (2.17-2.54)
R7P5
(4)
9.4*1.7
23.3*4.5
(21.0-30.1) (8.1-11.8)
13.9*3.0
(11.5-18.3)
1.49*0.19
2.89*0.59
(1.21-1.62) (2.52-3.76)
R2P5
(6)
23.5*2.8
9.3*1.3
(20.4-28.0) (7.8-11.1)
14.2*3.1
(11.1-19.9)
2.31*0.28
1.56*0.49
(1.11-2.46) (2.00-2.83)
R17P4
(6)
23.3*6.7
9.5*2.5
(15.2-32.8) (6.2-12.1)
13.8*4.4
(9.0-20.7)
1.45*0.17
2.22*0.35
(1.19-1.71) (1.71-2.67)
Average
(50)
22.2-5.1
8.9*2.0
13.3*3.6
1.50*0.31
2.38*0.40
Table 18.
Mean values and ranges among measurements for pachytene lengths and arm
ratios from nine tetraploid clones: Chromosome 9-D'.
Length (um)
Relative
Length
Clone
No.
Cells
Whole
Chromosome
Short arm
Long arm
B296
(5)
22.2-1.9
(20.1-24.2)
8.6-1.0
(7.1-9.6)
13.6*1.7
(II.O-I5.2)
1.60*0.27
2.54*0.32
(1.21-1.85) (2.28-3.03)
CLl
(2)
14.1-0.2
(13.9-14.2)
5.6*0.4
(5.3-5.9)
8.5*0.2
(8.3-8.6)
2.42*0.51
1.52*0.15
(1.41-1.62) (2.06-2.78)
Arm ratio
FBI41
(2)
19.4-1.2
(18.5-20.2)
7.1*1.5
(6.0-8.1)
12.3*0.3
(12.1-12.5)
2.22*0.12
1.79*0.42
(1.49-2.08) (2.13-2.30)
9946a
(7)
17.1*2.5
(13.5-20.6)
7.4*0.7
(6.1-8.1)
9.8*2.3
(7.4-13.6)
2.27*0.43
1.33*0.31
(1.14-1.94) (1.78-2.82)
FB49
(12)
22.9-5•1
9.1*1.7
(16.7-33.0) (6.6-12.6)
13.9*3.6
(9.1-20.4)
1.53*0.26
2.43*0.43
(1.12-1.85) (1.86-3.29)
R35P6
(6)
10.4*1.7
2 9 . 1* 3 . 7
(24.1-34.8) (8.6-12.7)
18.7*2.9
(15.4-22.8)
1.83*0.34
2.39*0.28
(1.39-2.42) (1.99-2.76)
R7P5
(4)
10.1*3.0
(8.2-14.5)
13.0*3.0
(11.1-17.5)
1.31*0.10
2.85*0.77
(1.21-1.44) (2.39-4.00)
R2P5
(6)
23.2-2.2
9.5*1.0
(20.9-27.2) (8.2-11.0)
13.7*2.5
(11.0-17.5)
2.30*0.31
1.47*0.35
(1.09-1.84) (1.99-2.75)
R17P4
(6)
24.3*6.0
9.7*2.6
(18.1-33.1) (6.6-13.4)
14.6*3.7
(10.5-19.7)
1.54*0.24
(1.12-1.74)
(2.03-2.69)
13.6*3.7
1.53*0.30
2.41*0.40
Average
(50)
23.0*6.0
(20.0-32.0)
22.5±5.3
8.9*2.0
2.33*0.22
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