CARYOLOGIAInternational Journal of Cytology, Cytosystematics

and Cytogenetics

Founded by ALBERTO CHIARUGI

ISSN 0008-7114

Published in Italyby

the University of Florence

April-June 2011Volume 64 - Number 2

*Corresponding author: e-mail koul_kk@yahoo.co.in

CARYOLOGIA Vol. 64, no. 2: 135-146, 2011

Spectrum of chromosomal confi gurations in pollen mother cells of Rhoeo spathacea (Swartz) Stearn

KOUL1,* KULDEEP KUMAR, RANJNA NAGPAL2 and ALOK ARUN1

1Department of Botany, Hindu College, Delhi-110007, India2Department of Botany, Ramjas College, Delhi-110007, India

Abstract — Rhoeo spathacea is unusual in that its twelve chromosomes are involved in complex translocations that results in atypical meiosis with all chromosomes frequently joined end-to-end into a single ring. The de-tailed meiotic studies, for the fi rst time, revealed that alignment of chromosomes into a ring and / or a chain begins at very early prophase I. However, the cells at diakinesis and metaphase I do not exclusively have a ring or a chain of 12 chromosomes. Instead, they carry a range of chain confi gurations formed due to slipping of the chiasmata off the pairing segments in complex structures. While in 57.61% cells, at diakinesis, a ring or a chain was observed, in the rest 2-7 chain confi gurations existed. Similarly, at metaphase I a ring of 12 chromosomes was observed in 33.87% cells whereas in 66.13 % cells 2-9 chain confi gurations existed. The various groups of chromosomes in cells at diakinesis generally did not show any specifi c orientation pattern. However, in 17.31% cells a disjunctional arrangement of chromosomes was observed. At metaphase I also while in 82.26% cells chromosomes showed more or less non disjunctional orientations, in 17.74% cells regular arrangement of chro-mosomes was observed. Owing to non-disjunctional arrangement of chromosomes at metaphase I, the anaphase I distribution of chromosomes was erratic in 44.18% cells. Three to nine chromosomes were seen at two poles in 33.06% cells whereas in 11.11% cells 1-8 lagging chromosomes existed. Regular distribution of 6:6 was seen in 55.82% cells. The Pollen sterility recorded was very high i.e. 81.24% using cotton blue staining procedure. It is suggested that a wide range of chromosome confi gurations, hitherto unidentifi ed, might exist in Rhoeo spathacea which awaits to be discovered.

Key words: chain confi gurations, non-disjunction, orientation, translocations.

INTRODUCTION

Rhoeo spathacea (Swartz) Stearn, also widely known as Rhoeo discolor Hance (family Com-melinaceae), is widely used as a favourite gar-den ornamental. This plant has interested the cytogeneticists all over the world ever since BELLING (1927) fi rst reported the presence of a ring of 12 chromosomes arranged end-to-end at diakinesis. Presence of such confi guration has long been interpreted as having resulted from extensive reciprocal translocations in which two arms of each chromosome synapse, respectively,

with an arm of two other chromosomes (BELL-ING 1927; DARLINGTON 1929; KATO 1930; SAX 1931; SIMMONDS 1945; LIN and PADDOCK 1973a; 1973b; 1978; VERMA and OHRI 1979; LIN 1980). Being an interesting material, extensive chromo-some studies have been reported by many au-thors (DARLINGTON 1929; KATO 1939; KOLLER 1932; SAX 1931, 1935; AKEMINE 1937; COLEMAN 1941; BHADURI 1942; SIMMONDS 1945; WALTERS and GERSTEL 1948; STEBBINS 1950; SWANSON 1957; WIMBER 1968; ZIMMERMAN 1968; MOENS 1972; NATRAJAN and NATRAJAN 1972; MERTENS 1973; LIN 1979a; 1979b; 1980; LIN and LEE 1979; VERMA and OHRI 1979; PETTANATI 1987; GOLCZYK and JOACHIMIAK 2003; GOLCZYK et al. 2005). However, it is disheartening to fi nd that inspite of all the earlier studies an insight into various meiotic confi gurations, particu-larly at early prophase I stage and metaphase I

KOUL, NAGPAL and ARUN136

stage, is far from satisfactory. Although KATO (1930) described the complete process of meio-sis using paraffi n sections in the typical form of Rhoeo, followed by studies carried out by LIN (1980; 1982), SATTERFIELD and MERTENS (1972), MERTENS (1973) and VERMA and OHRI (1979), the photomicrographs providing the details of range of confi gurations, particularly at metaphase I, have been more or less missing. The stickiness of chromosomes at early stages and extensive con-densation of chromosomes at metaphase I, that led to clump formation, has limited the detailed study of chromosome confi gurations (COLEMAN 1941; WALTERS and GERSTEL 1948; DESAI 1965; NATRAJAN and NATRAJAN 1972; VERMA and OHRI 1979). As far as we know most of the earlier information is mostly confi ned to the forma-tion of a ring confi guration and / or a few chain structures at diakinesis and comparatively little is known about meiotic confi gurations at met-aphase I stage.

The present study was initiated by our ob-servations of some new chromosomal confi gura-tions at both diakinesis and metaphase I stage while demonstrating meiosis to a group of un-dergraduates. This work puts on record the range of chromosome confi gurations seen at diakinesis and metaphase I of Rhoeo spathacea, supported by relevant photomicrographs, for the fi rst time.

MATERIAL AND METHOD

Meiosis was studied in Rhoeo spathacea grow-ing in the Botanic garden of Hindu College, Del-hi, India. The infl orescences of appropriate size were fi xed in acetic alcohol (1:3) for 24 hours and then preserved in 70% ethanol. For squash preparations anthers were taken from buds of proper size and squashed in 2% acetocarmine stain. All observations were made from tempo-rary slides. The photomicrographs were taken by using Olympus CH30 microscope. While making squash preparations care was taken to apply as little pressure as possible to avoid me-chanically distorting or breaking up of the chias-mata that held the chromosomes together. Only completely interpretable sets of chromosomes were scored both at diakinesis and metaphase I stage. Pollen viability was measured using the cotton blue staining procedure (SATTERFIELD and MERTENS 1972).

RESULTS

Rhoeo spathacea studied presently was a diploid with 2n = 12. At early prophase I the chromosomes appeared long thread like and instead of forming a densely tangled network of chromatin thread like structures, as is gen-erally observed, the chromatin threads were crowded together in a ring like fashion with a hollow clear centre (Figs. 1-2). While this was seen in 82% cells, in the remaining cells (18%) the tangled mass showed a linear / chain like arrangement.

At diplotene stage meiotic confi gurations ap-peared jumbled-up in a single clump though the ring like confi gurations could be seen. Diakine-sis was the most favourable stage available for study. Of the 387 pollen mother cells (PMCs) studied, while a single large ring of 12-chromo-somes was observed in 32.29% cells (Figs. 3-7), in the remaining 67.71% cells 1-7 chain confi gu-rations, containing varying number of chromo-somes, were observed (Figs. 8-30). Since the 12 chromosomes in a ring are held together pre-dominantly by the terminal chiasmata, a chain would be formed by the failure of one chiasmata between the pairing segment. Likewise 2,3,4 or more chain confi gurations with varying number of chromosome combinations would result from the failure of as many chiasmata and so on. Table 1 gives a concise account of the number and fre-quencies of various chromosomal associations observed in cells at diakinesis.

The chain confi gurations were found scat-tered all over in the cell. The cell with maximum number of chromosome groups (6-7 groups) had up to 4 univalents. While one chain of 12 chromosome was seen in 25.32% cells followed by 21.96% cells with 2 chain confi gurations, the cells with seven chains were infrequently observed (1.03%). In cells (32.29%) contain-ing one ring structure, the ring appeared too large to be accommodated in the nucleus with-out a certain amount of twisting and turning. The rings had 1-6 turns / twists (Figs. 3-7). In none of the cells was a fully opened out ring without any turn observed. Interestingly, while in 17.31% cells, carrying a ring or a chain of 12 chromosomes a zigzag (alternate) orientation of complex structures was observed (Fig. 8), in the rest the arrangement was random. However, in all the PMCs studied at diakinesis chromosomes were joined predominantly by terminal chias-mata though in 5 cells (1.29%) interstitial chias-mata were also observed.

MEIOTIC CONFIGURATIONS IN RHOEO SPATHACEA 137

Figs. 1-12 — Meiosis in Rhoeo spathacea. (1-2) PMCs at early prophase I. Note the arrangement of chromatin material. (3-7) Rings of 12 chromosomes at diakinesis showing varying twists and turns. (8) chromosomes showing alternate arrangement at diakinesis. (9-10) chain of 12 chromosomes at diaki-nesis. (11-12) cells at diakinesis showing 2 chains of 10+2 (Fig. 11) and 9+3 (Fig. 12). Bar = 10 µm.

KOUL, NAGPAL and ARUN138

At metaphase I, due to extreme condensation of chromosomes, the complex confi gurations ap-peared as clumps of varying sizes making the detailed analysis diffi cult. Nevertheless, a good

number of PMCs (124) were studied for chromo-some confi gurations and centromeric orientation (Figs. 31-52). Table 2 sums up the frequency of cells with different confi gurations at metaphase I.

Figs. 13-24 — PMCs at diakinesis in Rhoeo spathacea showing various chain confi gurations. (13) 5 + 3 + 2 + 2. (14) 5 + 3 + 2 + 1 +1. (15) 3 + 2 + 2 + 2 + 2 + 1. (16) 3 + 2 + 2 +2 + 1 + 1 +1. (17) 4 + 2 + 2 +1 +1 +1 +1. (18) 3 + 2 +2 + 2 +2 +1. (19) 5 + 2 +2 +1 +1 +1. (20) 4 + 3 + 2 + 1 +1 +1. (21) 6 + 5 + 1. (22) 7 + 4 +1. (23) 5 + 3 +3 +1. (24) 4 +3 + 2 +2 +1. Bar = 10 µm.

MEIOTIC CONFIGURATIONS IN RHOEO SPATHACEA 139

Of the 124 PMCs studied, while a ring of 12 chromosomes joined end-to-end was observed in 33.87% cells, a cell containing a chain of 12 chromosomes was never seen. The failure of chiasmata had resulted in breaking-up of ring structure into several confi gurations. In all 2-9 chain confi gurations were observed in 66.13% PMCs with higher number of combinations i.e. 8 and 9 observed only in 2.4l% cells. While in 17.74% PMCs disjunctional orientation of chro-mosome was observed (Fig. 33), in the rest al-though the chromosomes were condensed and

drawn towards the equatorial plate they did not show perfect disjunctional orientation; the con-fi gurations appeared both as zigzag and linear structures. The irregular arrangement of chro-mosomes was particularly evident in 15.3% cells with 2-8 univalents. It appeared each of the 12 chromosomes was equally likely to be involved in adjacent distribution regardless of the mor-phology of chromosome. For example, in a chain of four chromosomes while the two mid-dle chromosomes showed adjacent distribution and were destined to same pole, the two distal

Figs. 25-33 — PMCs of Rhoeo spathacea at diakinesis (25-30) and metaphase I (31-33), showing chro-mosome confi gurations of 3 + 3 + 2 + 2 + 2 (25); 7 + 5 (26); chain of 12 (27); 3 + 3 + 3 + 3 (28); 4 + 3 + 3 + 2 (29); 7 +5 (30); a ring of 12 chromosomes (31-32); 8 + 2 + 2 (33); note the alternate arrangement of chromosomes). Bar = 10µm.

KOUL, NAGPAL and ARUN140

chromosomes were destined for opposite pole. Similarly, while in a chain of fi ve chromosomes the middle two or three chromosomes showed adjacent arrangement, the chromosomes at ex-treme ends were facing the opposite pole. In a trivalent the chromosome were either facing the same pole or showed middle chromosome fac-ing one pole and the two distal chromosomes pointed towards the opposite pole. In occasion-al cells, with two chains of four chromosomes each, while one groups shows alternate arrange-ment the other had chromosomes with adjacent distribution. To be more precise, whatever has

been the number of confi gurations and combi-nations present in a cell, they never showed com-plete segregational orientations. This irregularity in chromosome orientation led to an abnormal numerical distribution of chromosome at ana-phase I. In the 369 PMCs studied while normal segregation of 6:6 was seen in 55.82% PMCs (Fig. 53), in the rest eleven different segregation patterns were observed (Figs. 54-60). Table 3 gives the frequency of cells with different types of distributions at anaphase I. While in 11.11% cells, 1-8 lagging chromosomes were observed with two poles receiving from one to seven chro-

TABLE 1 — Chromosome confi gurations in 387 of PMCs Rhoeo spathacea at diakinesis.

Confi gurations Chromosome combinations in each confi guration

(No. of PMCs)Total No.

of PMCs (%)Xta / cell (Xma

failure / cell)Total Xta (Total

Xma failure)One Ring 12(102). 225(32.29) 12(0) 1,500(0)

One Chain 12(98). 98(25.32) 11(1) 1,078(98)

Two Chains 11+1 (32); 10+2 (30); 9+3 (10); 8+4 (5); 7+5 (6); 6+ 6(2). 85(21.96) 10(2) 850(170)

Three Chains 10 +1+1(30); 9+2 +1(2); 8+3+1(2); 7 +4 +1(2); 6 + 5 +1(4).6+3+3 (2); 5 +4 (3).

45(11.62) 9(3) 405(135)

Four Chains 5+3+3 +1(4); 5 +3+2+2 (3); *5+4+2+1(2); 4+3+3+2(2);4+4+3+1(3); 3+3+3+3(2).

16(4.13) 8(4) 128(64)

Five Chains *5+3+2+1+1(2); 3+3+2+2+2 (1); 4+4+2+1+1 (3); 4+3+2+2+1(2).

8(2.06) 7(5) 56(40)

Six Chains *5+2+2+1+1+1(2); *4+3+2+1+1+1 (1); *3+2+2+2+2+1(2);4+2+2+1+1+1+1 (1).

6(1.55) 6(6) 36(36)

Seven Chains *3+2+2+2+1+1+1(4). 4(1.03) 5(7) 20(28)

* New reports.

TABLE 2 — Chromosome confi gurations in 124 PMCs at metaphase 1 of Rhoeo spathacea.

Confi gurations Chromosome combinations in each confi guration

(No. of PMCs)Total No. of

PMCs (% cells)Xta / cell (Xma

failure / cell)Total Xta (Total

Xma failure)One Ring 12(42) 42(33.87) 12(0) 504(0)

Two Chains 10+2(16); *9+3(10); 8+4(8). 34(27.41) 10(2) 340(68)

Three Chains *8+2+2(12); *7+3+2(4); *6+3+3(8). 24(19.35) 9(3) 216(72)

Four Chains 9+1+1+1(5); *7+2+2+1(5); *4+4+2+2(1). 11(8.87) 8(4) 88(44)

Five Chains *5+4+1+1+1(1); *4+3 + 3+1+1(1); 6+2+2+1+1(1);*4+4+2+1+1(1)

4(3.22) 7(5) 28(20)

Six Chains *4+2+2+2+1+1(2);*4+3+2+1+1+1(2); *3+3+2+2+1+1(2). 6(4.83) 6(6) 36(36)

Seven Chains *3+2+2+2+1+1+1(1). 1(0.80) 5(7) 5(7)

Eight Chains *5+1+1+1+1+1+1+1(1). 1(0.80) 4(8) 4(8)

Nine Chains *4+1+1+1+1+1+1+1+1(1). 1(0.80) 3(9) 3(9)

* New reports.

MEIOTIC CONFIGURATIONS IN RHOEO SPATHACEA 141

Figs. 34-51 — PMCs of Rhoeo spathacea at metaphase I with interpretive sketches showing chromosome confi gura-tions of 7 + 3 + 2 (34, 35); 3 + 2 + 2 +2 + 1 +1 +1 (36, 37); 9 + 1 +1 + 1 (38, 39); 9 +3 (40, 41); 5 + 4 + 1 +1 +1 (42, 43); 7 + 2 + 2 +1 (44, 45); 4 + 4 + 2 + 1 +1 (46, 47); 10 + 2 (48, 49); 4 + 3 + 2 +2 + 1 (50, 51). Bar = 10µm.

KOUL, NAGPAL and ARUN142

mosomes, in 33.06% cells, although lagging chromosome did not exist, varying chromosome numbers occurred at two poles i.e. 7:5, 8:4 and 9:3. The inversion bridges were observed in oc-casional cells (Fig. 59). The laggards observed at anaphase I were not included into the daugh-ter nuclei at telophase I indicating towards their loss or digestion by cytoplasmic enzymes.

The process of second meiosis was in general not unusual except that some microspores formed had hyper haploid (7,8) and / or sub haploid (5) chromosome number. At metaphase II the chro-mosomes were aligned at the equatorial plate with well spread out sister chromatids (Fig. 61). The anaphase II segregation was regular (Fig. 62).

The pollen viability was detected by cot-ton blue staining following SATTERFIELD and MERTENS (1972). Viable pollen (456; 18.76%) were uniform dark blue, while the nonviable pollen grains (1974; 81.24%) were atypical in shape, plasmolysed and vacuolated with no staining capacity (Figs. 63-64).

DISCUSSION

The present fi ndings are interesting on sever-al accounts. First, the arrangement of chromatin threads at early prophase I has been unlike the arrangement observed earlier. Second, a wide range of chromosome confi gurations observed in cells at diakinesis and metaphase I, followed by varying chromosomal segregation patterns at anaphase I, are being reported for the fi rst time.

At early prophase I the chromatin threads did not appear as a compact network. It formed a ring like structure in 82% cells with linear / chain like arrangement of chromatin threads vis-ible in 18% cells. It appears that the alignment of chromosomes into a ring and / or a chain con-fi guration starts at a very early stage. If we follow COLEMAN (1941), NATRAJAN and GROPP (1971) and NATRAJAN and NATRAJAN (1972), who at-tributed ring or chain formation to centromeric heterochromatin blocks which act as chromo-centres that drag in all chromosomes to initiate ring formation without showing any fusion and/or chiasmata formation, then one fi nds it easy to assign the early ring or chain formation to the in-volvement of heterochromatin blocks. But since no such heterochromatin blocks were studied it would be rather too premature to assign this reason. Nevertheless, the present observation of both a ring and / or a chain at very early stage could well be a pointer to the fact that the inter-

changed segment between the chromosomes re-sponsible for forming a ring i.e. 1eE and 12 de is too small which may or may not form a chiasma thus forming a ring or a chain. Here it will be appropriate to mention that in Rhoeo all the 12 chromosomes are arranged in a defi nite sequence and the two arms of each have been assigned the letters as 1eE-2ED-3Dc-4cC-5Cb-6bB-7Ba-8aA-9Af-10fF-11Fd-12de (SATTERFIELD and MERTENS 1972; LIN 1980). With this arrangement of chro-mosomes a ring would be formed only when chromosome 1eE and 12 de present at the two ends of the chain join. Earlier studies carried out at prophase I and particularly at pachytene stage have reported stickiness of chromosomes (COLEMAN 1941; WALTERS and GERSTEL 1948; DESAI 1965; NATRAJAN and NATRAJAN 1972 and VERMA and OHRI 1979) though KOLLER (1932) observed pairing between homologous segment of non-homologous chromosomes at pachytene stage. Owing to the stickiness of chromosome at early stages not much has been known about the behaviour of chromosomes at early stages. Whatever is known of the chromosome behav-iour with respect to paring and orientation it is mainly based on observations made at diakine-sis where interpretation of chromosome con-fi guration has been easy. At metaphase I too, barring the works of SATTERFIELD and MERTENS (1972), MERTENS (1973) and LIN (1980), not much is known regarding the confi gurations. The extreme condensation of chromosomes, that shrinks the complex confi gurations into clumped masses, have made the detailed analysis diffi cult (VERMA and OHRI 1979). Nevertheless, detailed analysis of chromosome confi gurations and orientations have been made in 369 and 124 PMCs studied at diakinesis and metaphase I, respectively. At both these stages a ring of 12-chromosomes and varying number of chain confi gurations, ranging from 1 to 7 (67.71%) at diakinesis and 2-9 (66.13%) per cell at met-aphase I, were commonly observed. While most of the chromosomal combinations observed at diakinesis confi rmed the earlier reports of SAT-TERFIED and MERTENS (1972), MERTENS (1973), VERMA and OHRI (1979) and LIN (1980), four chain combinations, carrying varying number of chromosomes in groups of 5 + 4 + 2 + 1, 3 + 3 + 3 +3, fi ve chains of 5 + 3 + 2 + 1 + 1 and six of 5 + 2 +2 + 1 + 1 + 1 and 3 + 2 + 2 + 2 + 2 + 1, hith-erto unreported, are being reported for the fi rst time. Similarly, in cells at metaphase I, 20 chro-mosome combinations were observed (Table 2) of which four combinations of 10 + 2, 8 + 4, 9

MEIOTIC CONFIGURATIONS IN RHOEO SPATHACEA 143

Figs. 52-64 — PMCs at metaphase I (52), anaphase I (53-60) and microspores at metaphase II and anaphase II (61-62), showing chromosome confi gurations of 5 + 1 + 1 + 1 + 1 + 1 + 1 +1 (52; note the arrangement of univalents) and segregation of 6: 6 (53); 5: 1: 6 (54); 7: 5 (55); 5: 4: 3 (56); 3: 8: 1 (57); 9: 3 (58); a bridge and a laggard (59); 5: 2: 5 (60). Microspore at metaphase II with 8 chromosomes (61) and anaphase II showing 6 chromatids at each pole (62). Figs. 63-64 show fertile and sterile pollen grains, respectively. Bar = 10µm.

KOUL, NAGPAL and ARUN144

+ 1 + 1 +1 and 6 + 2 + 2 + 1 + 1, besides a ring of 12 chromosomes, confi rmed the previous re-ports of VERMA and OHRI (1979) and LIN (1980) whereas the rest of the fi fteen combinations of 2-9 chain confi gurations (Table 2) are the fi rst re-port. Since all the chromosomes synapsed in one clump at early prophase I, the presence of vary-ing chromosome groups are the result of break-ing of complex structure once they were formed. One important observation that was particularly interesting and noticeable in cells at diakinesis and metaphase I was regarding the orientation of chromosomes in each group. While in most of the cells (82.69%) at diakinesis the different chromosome groups were randomly arranged, without showing any specifi c orientation pat-terns, in 17.31% cells the chromosomes showed disjunctional (i.e. alternate) orientation. At met-aphase I, although the chromosome were highly condensed and congressed at equatorial plate in majority of the cells, they showed both alternate and adjacent arrangement. The adjacent or non-disjunctional arrangement of chromosomes was particularly evident in cells with high univalent frequency. The frequency of cells showing regu-lar distribution, interestingly, was almost the same (17.74%), though marginally higher, with the frequency of cells at diakinesis i.e. 17.31%. The possible explanation might be that once the cells have assumed a stable confi guration at ear-lier stage they do not undergo further reorienta-tion at later stages. These observations not only are consistent with the observations made by NATRAJAN and NATRAJAN (1972) but also appear to support the concept of pre orientation ad-vocated by RICKARDS (1983) which implies that alternate and adjacent mode of orientation may drive, in part, directly from twisted and open ring confi gurations already present in diakinesis.

The non-disjunctional orientation of various groups of chromosomes and a very high number of independently oriented univalents resulted in discordant anaphase I segregation observed in 44.18% PMCs. These cells not only showed variable chromosome numbers at each pole (33.06%) but also had 1-8 lagging chromosomes in 11.11% PMCs. In all, twelve segregation pat-

terns were observed (Table 3) of which three patterns of 3:8:1, 5:6:1 and 5:5:2 were observed for the fi rst time. Owing to non-disjunctional arrangement a high frequency of sterile pollen grains (81.24%) were observed. Here it will not be out of place to mention that high pollen steril-ity has also been recorded in a number of other taxa like Petunia, Sorghum, Maize, Rice, Carrot, Onion and attributed to factors like a) meiotic irregularities which produce genetically unbal-anced pollen, b) irregularities in male gameto-phyte development and/or arrest of gameto-phyte development, c) premature dissolution of callose, d) viral/fungal infection, e) malfunction-ing of tapetum and/or hypertrophy of tapetal cells which invade the anther locule and crush the PMCs or pollen, and f) various environmen-tal factors like temperature (Oryza sativa ) and photoperiodic treatment (Glycine max, Silene pendula ) (see BHOJWANI and BHATNAGAR 1999). Pollen sterility has also been attributed to the presence of a recessive gene in the chromosome or a gene in the organelle other than nucleus. Be-sides the pollen sterility, ultrastructural changes in the pollen, before and after the anther dehis-cence, have also been studied in a number of angiosperms (SANGER and JACKSON 1971; BHO-JWANI and BHATNAGAR 1999). These include the structural changes in the cytoplasm (LARSON 1965), changes in generative cell and vegetative cell (ANGOLD 1968; BRIGHIGNA et al. 1981), the distribution of plastids (BRIGHIGNA and PAPINI 1997; MILOCANI et al. 2006), the degree of hydra-tion and the amount of soluble sugars (FRANCHI et al. 1996, 2002; MILOCANI et al. 2006). As far as Rhoeo is concerned, there is no information available on the changes in cytoplasm at ul-trastructural level during pollen development. However, as far as the pollen sterility or fertility is concerned, a perusal of literature available re-veals that in Rhoeo varying frequencies of pollen fertility have been recorded. For example, LIN and LEE (1979) observed 45.09% pollen fertility in a triploid Rhoeo whereas WALTER and GER-STEL (1948) reported 73% pollen fertility in a tetraploid cytotype. In diploid Rhoeo, while on one hand 70 to 90% pollen sterility has been re-

TABLE 3 — Chromosome segregation in 369 PMCs at anaphase I in Rhoeo spathacea.

Segregation 6:6 7:5 8:4 9:3 *3:8:1 7:1:4 5:4:3 6:2:4 5:1:6 *5:5:2 5:2:5 *5:6:1

No. of Cells 206 88 32 2 3 2 4 5 21 2 2 2

% Cells 55,82 23,84 8,67 0,54 0,81 0,54 1,08 1,35 5,69 0,54 0,54 0,54

* New reports.

MEIOTIC CONFIGURATIONS IN RHOEO SPATHACEA 145

corded (ZIMMERMANN 1968), on the other hand reports of very high fertility are also on record (KATO 1930; SAX 1931; AKEMINE 1937; BHADURI 1942; WALTERS and GERSTEL 1948; ZIMMERMAN 1968; LIN and PADDOCK 1973a, 1973b, MERTENS 1973; VERMA and OHRI 1979). Since the events of meiosis are genetically controlled it seems that in populations with high pollen fertility some strong selection for fertility, that tend to favour alternate orientation, is operational (LAWRENCE 1958; SUN and REES 1967; SYBENGA 1975; SYBEN-GA and RICKARDS 1987). However, with pollen sterility of upto 90% also on record one can not rule out the involvement of some unidentifi ed factor(s) that may have disturbed the genetic system inducing adjacent orientations.

Whatever be the reason for two extreme be-haviour of chromosomes with respect to orienta-tions, it is clear that non disjunctional orienta-tion have led to high pollen sterility. This view is strongly supported by the fact that despite ob-serving regular segregation of 6:6 chromosomes in 55.82% PMCs, where one would not expect less than 50% pollen fertility, a high pollen ste-rility was recorded. It seems probable that these cells, though receiving normal chromosome number, carried disharmonious genetic comple-ment owing to adjacent orientation of chromo-somes at metaphase I.

In conclusion, Rhoeo spathacea represents an example of a complex translocation hetero-zygote. The 12-chromosomes do not exclusively form the ring structure, instead an array of chro-mosome confi gurations occur besides the ring confi gurations at both diakinesis and metaphase I. These confi gurations mostly show non dis-junctional orientation leading to high pollen ste-rility. Presence of a wide range of confi gurations suggest that the discovery of various infrequent chromosome confi gurations only awaits study of suffi ciently large number of cells.

Acknowledgement — The help provided by Dr. A.C. Handa of Chemistry Department, Hindu Col-lege during photography is thankfully acknowledged.

REFERENCES

ANGOLD R.E., 1968 — The formation of the generative cell in the pollen grains of Endymion non-scriptus. Journal of Cell Science, 3: 573-578.

AKEMINE T., 1937 — Non-disjunction of the meiotic chromosomes in Rhoeo discolor Hance. Japanese Journal of Genetics, 13; 31-36.

BELLING J., 1927 — The attachment of chromosomes

at the reduction division in fl owering plants. Jour-nal of Genetics, 18: 177-205.

BHADURI P.N., 1942 — Cytological analysis of struc-tural hybridity in Rhoeo discolor Hance. Journal of Genetics, 44: 73-85.

BHOJWANI S.S. and BHATNAGAR S.P., 1999 — The Em-bryology of Angiosperm. Vikas publishing House Pvt. Ltd., New Delhi, India.

BRIGHIGNA L, A. CECCHI-FIORDI and PALANDRI M.R., 1981 — Ultrastructural investigation on two-nucle-ate pollen grain of Tillandsia caput-medusae Morr. American Journal of Botany, 68(8): 1033-1041.

BRIGHIGNA L. and PAPINI A., 1997 — Plastidial dy-namics during pollen development in Tillandsia al-bida Mez & Purpus (Bromeliaceae) before anthesis. Phytomorphology, 47(1): 59-65.

COLEMAN L.C., 1941 — The relation of chromocent-ers to the differential segments in Rhoeo discolor Hance. American Journal of Botany, 28: 742-748.

DARLINGTON C.D., 1929 — Chromosome behavior and structural hybridity in the Tradescantiae. Jour-nal of Genetics, 21: 207-286.

DESAI S.A., 1965 — Cytological study of a tertaploid Rhoeo discolor. Cytologia, 30: 260-265.

FRANCHI G.G., BELLANI L., NEPI M. and PACINI E., 1996 — Types of carbohydrates reserves in pollen: localization, systematic distribution and ecophysi-ological signifi cance. Flora, 191: 1-17.

FRANCHI G.G., NEPI M., DAFNI A. and PACINI E., 2002 — Partially hydrated pollen: taxonomic dis-tribution, ecological and evolutionary signifi cance. Plant Systematics and Evolution, 234: 211-227.

GOLCZYK H., HASTEROK R. and JOACHIMIAK A.J., 2005 — FISH-aimed karyotyping and characterization of Renner complexes in permanent heterozygote Rhoeo spathacea. Genome, 48: 145-153.

GOLCZYK H. and JOACHIMIAK A., 2003 — NORs in Rhoeo (Commelinaceae) revisited. Caryologia, 56(1): 31-35.

KATO K., 1930 — Cytological studies of pollen mother cells of Rhoeo discolor Hance, with special refer-ence to the question of the mode of synthesis. Me-morial College of. Science. Kyoto Imperial Uni-versity Series B., 5: 139-161.

KOLLER P., 1932 — Further studies in Tradescantia vir-giniana var. humilis and Rhoeo discolor Journal of Genetics, 26: 81-96.

LARSON D.A., 1965 — Fine structural changes in the cytoplasm of germination pollen. American Jour-nal of Botany, 52: 139-154.

LIN Y.Z., 1979a — Chromosome distribution and cat-enation in Rhoeo spathacea var. concolor. Chromo-soma, 71: 109-127.

LIN Y.Z., 1979b — Fine structure of meiotic prophase chromosomes and modifi ed synaptonemal complex-es in diploid and triploid Rhoeo spathacea. Journal of Cell Science, 37: 69-84.

LIN Y.J., 1980 — Chromosome behaviour in Rhoeo spathacea var. variegata. Cytobios, 27: 113-128.

LIN Y.J., 1982 — Chiasma failures and chromosome association in Rhoeo spathacea var. variegata Cyto-bios, 33: 7-14.

KOUL, NAGPAL and ARUN146

LIN Y.J. and LEE I.T., 1979 — Studies of Chromosome distribution in a triploid Rhoeo. Caryologia, 32(2): 205-214.

MERTENS T.R., 1973 — Meiotic chromosome behavior in Rhoeo spathacea. Journal of Heredity, 64: 365-368.

MILOCANI E., PAPINI A. and BRIGHIGNA L., 2006 — Ultrastructural studies on bicelluler pollen grains of Tillandsia seleriana Mez (Bromeliaceae), a neo-tropical epiphyte. Caryologia, 59(1): 88-97.

MOENS P.B., 1972 — Fine structure of chromosome coiling at meiotic prophase in Rhoeo discolor. Ca-nadian Journal of Genetics and Cytology, 14: 801-805.

NATRAJAN A.T. and GROPP G., 1971 — The meiotic behaviour of autosomal heterochromatin segments in hedgehogs. Chromosoma, 35: 143-152.

NATRAJAN A.T. and NATRAJAN S., 1972 — The hetero-chromatin of Rhoeo discolor. Hereditas, 72: 323-330.

PETTENATI M.J, 1987 — Giemsa C-banding of Rhoeo (Commelinaceae). Genetica, 74: 219-224.

RICKARDS G.K., 1983 — Orientation behaviour of chromosome multiples of interchange (reciprocal translocation) heterozygotes. Annual Review of Genetics, 17: 443-498.

SANGER J.M. and JACKSON W.T., 1971 — Fine structure study of pollen development in Haemanthuskath-erinae Barker. II. Microtubule and elongation of generative cells. Journal of cell Science, 8: 30-315.

SAX K., 1931 — Chromosome ring formation in Rhoeo discolor. Cytologia, 3: 36-53.

SAX K., 1935 — Chromosome structure in the meiotic chromosomes of Rhoeo discolor Hance. Journal of Arnold Arboretum, 16: 216-224.

SIMMONDS N.W., 1945 — Meiosis in tropical Rhoeo

discolor. Nature, 155: 731.SATTERFIELD S.K. and MERTENS T.R., 1972 — Rhoeo

spathacea: A tool for teaching meiosis and mitosis. Journal of Heredity, 63: 375-378.

STEBBINS G.L., 1950 — Variation and evolution in higher plants. Columbia University Press, New York, 643 pp.

SUN S. and REES H., 1967 — Genetic control of chro-mosome behaviour in rye IX. The effect of selection on the disjunction frequency of interchange associa-tions. Heredity, 22: 249-254.

SWANSON C.P., 1957 — Cytology and Cytogenetics. MacMillan and Co. Limited, Englewood, New Jersey.

SYBENGA J., 1975 — Meiotic confi gurations. Springer Verlag, Berlin, Heidelberg, New York.

SYBENGA J. and RICKARDS G.K., 1987 — The orienta-tion of multivalents at meiotic metaphase I: a work-shop report. Genome, 29: 612-620.

VERMA S.C. and OHRI D., 1979 — Breakdown of the classical meiotic system in Rhoeo spathacea (Com-melinaceae). Cytologia, 44: 91-102.

WALTERS M.S. and GERSTEL D.U., 1948 — A cyto-logical investigation of a tetraploid Rhoeo discolor. Americal Journal of Botany, 35: 141-150.

WIMBER D.E., 1968 — The nuclear cytology of biva-lent and ring forming Rhoeos and their hybrids . American Journal of Botany, 55: 572-574.

ZIMMERMAN E., 1968 — Mechanism of non disjunc-tional chromosome distribution and the cause of pollen sterility in Rhoeo spathacea. Chromosoma, 25: 215-248.

Received June 3rd 2010; accepted March 7th 2011