Phaseolus Mungo L. (Synonym: Vigna Mungo (L.) Hepper.) Is Black Gram
Total Page：16
File Type：pdf, Size：1020Kb
Cytologia 40, 787-795 1975 Black Gram•~Rice Bean Cross Majibar R. Biswas and S. Dana Department of Genetics and Plant Breeding , University of Kalyani, Kalyani 741235, Nadia, West Bengal , India Received June 21, 1974 Phaseolus mungo L. (synonym: Vigna mungo (L .) Hepper.) is black gram, a pulse crop widely cultivated in India. Phaseolus calcaratus Roxb. (synonym: Vigna umbellata (Thunb.) Ohwi and Ohashi) is known as rice bean and is cultivated in India both as pulse and fodder crop . Reports on crosses of P. mungo with P . aureus, P. trilobus and tetraploid Phaseolus species and of P . calcaratus with P. aureus and tetraploid Phaseolus species have indicated that these species are isolated from each other by hybrid inviability and weakness and hybrid sterility types of isolating barriers (Dana 1965a, b, 1966 a, b, c, 1967, 1968). The present work at tempts to understand the nature of isolating mechanisms particularly the hybrid sterility operating between these two species. Materials and methods The seeds of P. mungo variety T. 9 were collected from Economic Botanist, Uttar Pradesh, Kanpur; variety N. P. 14 from Division of Genetics, Indian Agri cultural Research Institute, New Delhi and variety W. B. 16 from Economic Bo tanist, West Bengal. The seeds of P. calcaratus (accession no. EC 18183) were obtained from Division of Plant Introduction, Indian Agricultural Research Insti tute, New Delhi. The techniques of hybridization and pollen fertility analysis followed here were same as those described by Dana (1965a). Cytological preparations were made according to the method described by Dana (1966d). Germination of hybrid seeds: The shrivelled or partially developed mature hybrid seeds were surface sterilized with absolute ethyl alcohol and soaked for 24 hours in sterilized nutrient solution of Sanders, Franzke and Ross (1959) as modified by Dana (1964). The embryonic axes were dissected out of the soaked seeds and were placed on cotton wads soaked in the nutrient solution with 4 percent sucrose. The cotton wads were kept within petridishes. The petridishes along with soaked cotton wads were previously sterilized by autoclaving at 15 lbs pressure for 30 minutes. The dissection and transfer of embryonic axes were done under aseptic conditions. The petridishes containing the embryonic axes were kept in diffuse light for 4 days. The solution within the petridishes was replaced every day by fresh, sterilized culture solution. After this period, the growing seedlings were transplanted in earthen pots (30cm diameter) containing soil and leaf compost in the proportion of 1: 1 by volume. The method of colchicine treatment of seedlings was same as that described by 788 M. R. Biswas and S. Dana Cytologia 40 Sen and Chheda (1958). Observations Crossability Three varieties of P. inungo-N. P. 14, W. B. 16 and T. 9 and only one variety of P. calcaratus (EC 18183) were used in hybridization. The cross between these two species succeeded only when P. mungo was used as female parent. In P. calcaratus •Š•~P. mungo •‰ crosses, a total of 116 pollinations were made but all the crossed pods shed within 2 weeks. In P. mungo (variety N. P. 14) •ŠxP. calcara tus •‰ cross, 48 pollinations were made of which only 3 pods reached maturity and these pods contained 7 shrivelled and 4 empty seeds. In P. mungo (variety W. B. 16) •Š•~P. calcaratus •‰ cross, 35 pollinations produced only 2 pods containing 6 shrivelled seeds. All the, seeds of these two crosses did not germinate. In P. mungo (variety T. 9) •Š•~P. calcaratus •‰ cross, 240 pollinations resulted in 48 crossed pods containing 145 shrivelled and 9 empty seeds. All these empty seeds and 110 shrivelled seeds did not germinate. The rest 35 shrivelled seeds of this cross were cultured in nutrient solution with sucrose. Out of these only 29 germinated, 22 seedlings survived for a week and 12 plants reached flowering stage. F1 generation The hybridity of F1 seedlings was confirmed by their hypogeal germination which was a characteristic of the male parent, P. calcaratus. The hybrids were weak and stunted in growth during the winter but they put up vigorous growth in summer. The general growth habit of the parents and F1 is shown in Figs. 1, 2 and 3. The chara cters of P. mungo dominating in F1 were colour of stem and margin of leaflets (Fig. 4). The hypogeal germination and general growth habit of P. calcaratus parent were dominant in F1. The hybrid was intermediate between the two parents in colour of petiole and most of the metrical characters at flowering stage. The characters of the parents and F1 are compared in Table 1. The plants of P. mungo (variety T. 9) and P. calcaratus had on an average 98.5 and 97.2 per cent pollen fertility respectively whereas the F1 plant had only 0.7 per cent fertile pollen. In F1, all the flowers shed within 48 hours after opening and thus no pod set was noted. The two parental species and the F1 had 2n=22 chromosomes. Meiosis in both the parental species was normal with regularly formed 11 bivalents. Meiosis in F1 was highly irregular. In F1, a total of 59 cells were analysed at MI and complete bivalent formation was not observed in any one of these cells. Highest number of bivalents noted in a cell was seven and only 11.9 per cent of cells showed this con figuration. Both 6 bivalents with 10 univalents and 3 bivalents with 16 univalents were observed in maximum number of cells i. e. 18.6 per cent. One quadrivalent was noted in a total of 5.1 per cent of cells (Fig. 5). In rest of the cells 5, 4, 2 and 1 bivalents with 12, 14, 18 and 20 univalents respectively were observed (Figs. 6, 7). In most of the cells lack of coorientation of bivalents on metaphase plates and precocious movement of the univalents were noted (Figs. 6, 7). The average chromo Black Gram •~ Rice Bean Cross 789 Figs. 1-4. Photographs of plants and leaves. 1, P. mungo (var. T. 9.). 2, P. calcaratus (var. E. C. 18183). 3, F1 P. mungo •~P. calcaratus, 4, terminal leaflets of parents and Fl. •~1/2 nat. size. 790 M. R. Biswas and S. Dana Cytologia 40 Table 1. Comparison of parents and F1 Black Gram •~ Rice Bean Cross 791 some configuration per cell in F1 was 0.05IV(range 0-1)+4 .24II (range 0-7)+13.32, (range 8-20). Out of 91 cells analysed at Al, only 8.8 per cent had normal 11/11 segregation. Dicentric bridges with acentric fragments were observed in 3.3 per cent of cells (Fig. 8). In rest of the cells, 2 to 12 laggards were noted . Spindle abnormalities were noticed in majority of the cells (Fig. 8). Figs. 5-11. Photomicrographs of meiosis in F1. 5, 1 IV(•¨)+5 II+8I at MI. •~1100. 6, MI with 4II+14I. One I (•¨) overlaped with a II. •~2000. 7, 2 I1+18I at MI. •~1500. 8, Al showing dicentric bridge with fragment. •~1100. 9, A II showing dicentric bridge. •~1100. 10, multiple poles at All. •~1500. 11, univalent (•¨),loosely paired bivalents (•¨•¨) and differential segments in bivalents (+•¨) at pachytene. •~1800. 792 M. R. Biswas and S. Dana Unequal segregation of chromosomes and spindle abnormalities were also evi dent at All. A total of 42 cells were analysed of which only 4.8 per cent of cells showed equal segregation of chromosomes. Tripolar movement and dicentric bridges were noted in 19.0 and 4.7 per cent of cells respectively (Fig. 9). In rest of the cells unequal separation with laggards and movement of chromosomes to 5 to 8 poles were observed (Fig. 10). At pachytene stage in Fl, the pairing of chromosomes was not complete. Many univalents and some loosely paired bivalents were observed in most of the cells. In many of these bivalents terminal and/or interstitial segments remained unpaired (Fig. 11). Amphidiploid Four hybrid seedlings at the age of 5 days were treated with 0.5per cent coichi cine in 0.5 per cent agar medium for 6 hours daily for 3 consecutive days. Two of these treated seedlings died within 7 days after treatment and the other 2 seedlings survived upto flowering. The treated plants had stunted growth for 15 days after which they showed more vigorous growth. These plants had on an average 81.0 per cent pollen fertility and the average diameter of fertile pollen was 54.5ƒÊ. One of these treated plants produced 3 pods and the other only 1 pod. From these 4 pods, 2 partially filled and 4 well developed seeds were obtained. As the F, was sterile, the pod and seed characters of amphidiploids i.e. colchi cine induced fertile Fl plants were compared with those of the parents. The amphi diploids were like P. mungo parent in the position of pods on peduncle, tip of the young pods and hilum of the seeds. They were like P. calcaratus parent in the sur face of pods and were intermediate between the two parents in colour of mature pods and colour of seed coat (Table 2). Table 2. Comparison of parents and amphidiploid Discussion The two species involved in the present cross are separated from each other, besides their self-pollinating device, by post fertilization barriers like hybrid inviabi Black Gram x Rice Bean Cross 793 lity, weakness and sterility. Hybrid inviability which is the immediate result of incompatibility between embryo and endosperm, have led to the formation of shrivel led and empty hybrid seeds in the present study. The empty seeds were inviable .