ISSN 09734031 Genetics of fertility restoration of Wild Abortive cytoplasmic male sterility in rice (Oryza sativa L.) S. Sreedhar *, T. Dayakar Reddy 1 and M.S Ramesha 2 Department of Genetics and Plant breeding, College of Agriculture, ANGRAU, Rajendranagar, Hyderabad500030 (A.P.), India 1 Professor, College of Agriculture, ANGRAU, Rajendranagar, Hyderabad500030, India 2 Rice scientist, CSISA, IRRI, Philippines *E mail: siddu.35@gmail.com ABSTRACT Genetics of fertility restoration in four restorer lines viz., IR697025233R, IR62161 1893132R, KBNT11 and IR596699313R was studied in Wild Abortive cytoplasmic male sterility system using cytoplasmic male sterile line, APMS6A. It was found that fertility restoration evaluation of the plants in F 2 of all the four crosses viz., APMS 6A x IR69702523 3R, APMS 6A x IR621611893132R, APMS 6A x KBNT11 and APMS 6A x IR5966993 13R showed a segregating ratio of 15 fertile: 1 sterile indicated digenic inheritance with dominant duplicate gene action. It can be concluded that the presence of dominant duplicate genes in four restorer lines viz., IR697025233R, IR621611893132R, KBNT11 and IR 596699313R governing fertility restoration in the WACMS system. KEY WORDS: Dominant duplicate gene action, fertility restoration, genetics, Oryza sativa, wild abortive INTRODUCTION Rice is one of the most important cereal crops in India. Spectacular growth in rice production was achieved since 1960s through the adoption of semidwarf varieties coupled with other green revolution technologies that made the country selfsufficient in rice during 1980s. However, the productivity has come to stagnation since last two decades and efforts have failed to give tangible results to break the genetic yield barrier in rice. To make the India selfsufficient in rice and to meet future food requirements, it is needed to improve the productivity every year by almost two million tonnes. The task is quite challenging, in view of plateauing trend observed in yield potential of high yielding varieties, shrinking land, dwindling water resources and nonavailability and sky rocketing costs of labour, and at the same time preserving the quality of environment. Hybrid rice technology is a key strategy for increasing rice production and addressing food security and rice selfsufficiency. Among the many available genetic approaches being exploited to break the yield barrier in rice, hybrid rice technology appears to be the most appropriate, ecofriendly and readily adoptable one. Results of the hybrid rice commercialization program in the China look promising. China has been able to 412
ISSN 09734031 produce 200 million tonnes rice annually from 30 million hectares. It has been also demonstrated in India very clearly on large scale that hybrids give 1520 per cent increased yield over the highest yielding varieties under the similar growing conditions by using Wild Abortive (WA) cytoplasmic male sterility. The discovery of cytoplasmic male sterility (CMS) not only facilitates hybrid seed production, but also provides an excellent system for the study of nucleuscytoplasm interaction. High yield potential of CMS derived F 1 hybrids depends upon their high pollen fertility and spikelet fertility, which determined by the number and mode of action of restorer genes present in their restorer parent. Knowledge of the genetic control of male fertility restoration and extent of fertility restoration facilitates, transfer of fertility restorer genes to promising breeding lines and undertake improved restorer breeding programme and ultimately their deployment in hybrid breeding programme. In the present investigation, involving four crosses viz., APMS 6A x IR697025233R, APMS 6A x IR621611893132R, APMS 6A x KBNT11 and APMS 6A x IR 596699313R was undertaken to understand the genetics of fertility restoration of CMS line of WA cytoplasm by four crosses. MATERIAL AND METHODS In the present investigation, one CMS line, APMS 6A and four restorer lines, IR697025233R, IR621611893132R, KBNT11 and IR596699313R were used in the study is listed in Table 1. The CMS line, APMS6A inherited WA cytoplasm imparting male sterility. The restorer lines, IR697025233R, IR621611893132R, KBNT11 and IR596699313R were crossed with the cytoplasmic male sterile line, APMS6A during kharif, 2009. The four fertile F 1 s viz., APMS 6A x IR69702523 3R, APMS 6A x IR621611893132R, APMS 6A x KBNT11 and APMS 6A x IR 596699313R were selfed to raise the F 2 generation. Twenty five grams of F 2 seed each of the entire four cross combinations were sown and more than 1000 population of F 2 was raised. Seedlings of the parental, F 1 and F 2 were transplanted 20 x15 cm apart using single seedling per hill. Two panicles of each plant in parental, F 1 and F 2 were bagged at panicle emergence to avoid out crossing. The bagged at panicles of individual plants were separately harvested at maturity and fertile and sterile spikelets were counted per cent spikelet fertility was worked out. The F 2 progeny of four cross combinations viz., APMS 6A x IR69702 5233R, APMS 6A x IR62161189313 2R, APMS 6A x KBNT11 and APMS 6A x IR596699313R, along with their parents and F 1 s were evaluated during rabi, 200910 at Directorate of Rice Research (DRR), Hyderabad. Pollen fertility and seedsetting rate were used as the main criteria for the evaluation of fertile and sterile plants. Mature anthers were harvested, and the pollen was stained with 1% iodine in potassium iodide solution. The number of dark blue (stainable) and clear pollen grains (unstainable) in each individual were counted under an optical microscope (Plate 1) and the numbers of the seed set on a panicle were counted. Classification of F 2 population based on pollen fertility and spikelet fertility in to following classes (Table 2): 413
ISSN 09734031 Fertile: Plants showing more than 80 percent pollen fertility and more than 75 per cent spikelet fertility. Partially fertile: Plants showing 50.1 to 80 per cent pollen fertility and 50.1 to 75 per cent spikelet fertility. hybrids dehisced normally and more than 80 per cent of pollen was stained in all the four hybrids and spikelet fertility of F 1 hybrids was more than 90 per cent in all the four crosses studied indicating that fertility restoration in the relevant pollinators is under dominant gene control. Partially sterile: Plants showing 1.1 to 50 per cent pollen fertility and 0.1 to 50 per cent spikelet fertility. Sterile: Plants showing zero to 1 per cent pollen fertility and zero per cent spikelet fertility. The F 2 plants of cross combinations were grouped into two classes a plant with 01 per cent pollen fertility and zero per cent seed set as sterile and a plant with above one per cent pollen fertility and above zero per cent seed set as fertile, considering that sterile or fertile has the characteristic of a qualitative character. For chisquare analysis, fertile, partially fertile and partially sterile plants were grouped into one class of male fertile plants clearly distinguished from male sterile plants that did not shed any pollen. The data, thus generated, were subjected to chisquare (χ 2 ) test. Chisquare is a test of significance to test the goodness of fit between observed values and expected values based on hypothesis. RESULTS AND DISCUSSION In the present investigation, the results indicated that the CMS line, APMS 6A was completely sterile, whereas the pollinators, IR697025233R, IR62161 1893132R, KBNT11 and IR5966993 13R were fully fertile. The anthers of the Segregation analysis of the fertility of F 2 plants of the hybrid combinations was made to determine the number of genes controlling sterility or fertility. Segregation pattern for pollen fertility and spikelet fertility in F 2 generation of the four hybrid combinations viz., APMS 6A x IR69702 5233R, APMS 6A x IR62161189313 2R, APMS 6A x KBNT11 and APMS 6A x IR596699313R along with their F 1 s and parents is presented in Table 3. The results further revealed that F 2 population of all the four crosses viz., APMS 6A x IR697025233R, APMS 6A x IR 621611893132R, APMS 6A x KBNT 11 and APMS 6A x IR596699313R exhibited a segregating ratio of 15 fertile: 1 sterile, since calculated χ 2 values were found to be lower than the table value of chisquare (3.84, at p = 0.05) indicated digenic inheritance with dominant duplicate gene action governing fertility restoration in the WACMS system. The chisquare test confirmed the goodness of fit of 15 fertile: 1 sterile in all the four crosses studied. This indicated the presence of dominant duplicate genes in IR69702 5233R, IR621611893132R, KBNT 11 and IR596699313R governing fertility restoration in the WACMS. The literature collected indicated that the inheritance of fertility restoration of WA 414
ISSN 09734031 cytoplasm is under the control of single, two, three or even four genes and mode of action may vary with the restorers used. Majority of the previous reports (Bharaj et al.,1991; Ramalingam et al.,1992; Singh et al.,1994; Bharaj et al.,1995; Virmani et al.,1997; Sridhara et al.,1998; Pradhan and Jachuk,1999; Sharma et al., 2001, Yograj et al.,2002; Sharma and Singh, 2003; Haohua et al.,2006; Tan et al.,2008; Sattary et al., 2008 and Sheeba et al.,2009) suggest that the fertility restoration of WACMS lines was under the control of two independently segregating dominant genes. The effect of one of the two genes in restoring fertility appears to be stronger than the other, because the presence of one of the genes alone conferred partial fertility (Bharaj et al., 1991; Bharaj et al., 1995; Virmani et al., 1997; Yograj et al., 2002 and Sharma and Singh, 2003). Interestingly in the present investigation, the mode of action of the two genes was duplicate gene action in all the four crosses studied i.e. epistasis with duplicate dominance. Reports of similar type of gene interaction were put forth by Math et al., 1990; Pande et al., 1990; Sridhara et al., 1998; Yogaraj et al., 2002; Sattary et al., 2008 and Sheeba et al., 2009. The use of restorers with more than one independent gene for fertility restoration is likely to produce the hybrids with higher fertility restoration and consequently increased yield. The identified restorers would be useful in future hybrid breeding programmes to develop more restorers with diverse genetic background. Table 1: List of male sterile lines and restorer lines used in the study S.No. Genotype Source CMS / Restorer line 1 APMS 6A RARS, Maruteru 2 IR697025233R IRRI, Philippines 3 IR621611893132R IRRI, Philippines 4 KBNT11 IRRI, Philippines 5 IR596699313R IRRI, Philippines Table 2: Classification of F 2 population based on pollen fertility and spikelet fertility Class Pollen fertility per cent Spikelet fertility per cent Fertile > 80 > 75 Partially fertile 50.180 50.175 Partially sterile 1.150 0.150 Sterile 01 0 415
ISSN 09734031 Table 3: Segregation pattern of male fertile (F) and male sterile (S) plants in P 1, P 2, F 1 and F 2 populations of four crosses in rice Genera tions Total plants Observed frequencies Fertile (F) Sterile (S) Expected frequencies Fertile Sterile (F) (S) APMS 6A x IR697025233R P 1 48 48 P 2 42 42 F 1 32 32 F 2 842 790 52 789.38 52.63 APMS 6A x IR621611893132R P 1 48 48 P 2 46 46 F 1 46 46 F 2 868 813 55 813.75 54.25 APMS 6A x KBNT11 P 1 48 48 P 2 43 43 F 1 40 40 F 2 908 851 57 851.25 56.75 APMS 6A x IR 596699313R P 1 48 48 P 2 45 45 F 1 41 41 F 2 1017 954 63 953.44 63.56 Ratio F:S χ 2 χ 2 table value 15:1 0.007 3.841 15:1 0.011 3.841 15:1 0.001 3.841 15:1 0.004 3.841 416
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