Cytoplasmic-nuclear Male Sterility Research Articles

Cytoplasmic genome contributions to domestication and improvement of modern maize

BackgroundStudies on maize evolution and domestication are largely limited to the nuclear genomes, and the contribution of cytoplasmic genomes to selection and domestication of modern maize remains elusive. Maize cytoplasmic genomes have been classified into fertile (NA and NB) and cytoplasmic-nuclear male-sterility (CMS-S, CMS-C, and CMS-T) groups, but their contributions to modern maize breeding have not been systematically investigated.ResultsHere we report co-selection and convergent evolution between nuclear and cytoplasmic genomes by analyzing whole genome sequencing data of 630 maize accessions modern maize and its relatives, including 24 fully assembled mitochondrial and chloroplast genomes. We show that the NB cytotype is associated with the expansion of modern maize to North America, gradually replaces the fertile NA cytotype probably through unequal division, and predominates in over 90% of modern elite inbred lines. The mode of cytoplasmic evolution is increased nucleotypic diversity among the genes involved in photosynthesis and energy metabolism, which are driven by selection and domestication. Furthermore, genome-wide association study reveals correlation of cytoplasmic nucleotypic variation with key agronomic and reproductive traits accompanied with the diversification of the nuclear genomes.ConclusionsOur results indicate convergent evolution between cytoplasmic and nuclear genomes during maize domestication and breeding. These new insights into the important roles of mitochondrial and chloroplast genomes in maize domestication and improvement should help select elite inbred lines to improve yield stability and crop resilience of maize hybrids.

Inheritance Pattern of A1 Cytoplasmic-nuclear Male Sterility System in Commercial Pearl Millet Hybrids [Pennisetum glaucum (L). R. Br.] in Summer and Post-rainy Seasons

Background: This study aimed to understand the genetic aspects and inheritance patterns that control the process of restoring fertility in the context of the A1 system of Cytoplasmic-Nuclear Male Sterility (CMS) in pearl millet. Methods: The experimentation was conducted in commercial pearl millet hybrids, utilizing three F2 populations and three BC1 populations. These populations underwent assessment during the rainy and post-rainy seasons at Rallis India Limited, Hyderabad During the Post-rainy (November-February) and summer (March-June) seasons of 2022-2023. Segregation data derived from observations on pollen shedding and seed set were gathered and subsequently, statistical analysis was performed. The Chi-square (χ2) test was employed to assess the conformity of the observed segregation data to the expected genetic ratios, thus evaluating the goodness of fit. Result: The hybrid combinations MP13P025A ×MP16P050R (MP7171) and MP18P069A × MP20P081R (MP7214) consistently displayed a 3:1 segregation ratio in the F2 generation and a 1:1 ratio in the BC1 generation, regardless of the rainy or post-rainy seasons. However, the cross between MP14030A and MP15P038R (MP7878) showed irregularities in both seasons due to modifiers and environmental factors. These irregularities stemmed from an excess of fertile plants (FF) in the post-rainy season or an excess of sterile plants (FS) in the summer season. These observations provide valuable insights into the genetic dynamics of the A1 Cytoplasmic-Nuclear Male Sterility (CMS) system within commercial pearl millet hybrids. The study indicates that fertility restoration in the A1-based CMS system is strongly influenced by a single Dominant gene control mechanism governing the interplay between male sterility and fertility restoration in these crosses.

Genomic prediction of hybrid performance for agronomic traits in sorghum.

Hybrid breeding in sorghum [Sorghum bicolor (L.) Moench] utilizes the cytoplasmic-nuclear male sterility (CMS) system for seed production and subsequently harnesses heterosis. Since the cost of developing and evaluating inbred and hybrid lines in the CMS system is costly and time-consuming, genomic prediction of parental lines and hybrids is based on genetic data genotype. We generated 602 hybrids by crossing two female (A) lines with 301 diverse and elite male (R) lines from the sorghum association panel and collected phenotypic data for agronomic traits over two years. We genotyped the inbred parents using whole genome resequencing and used 2,687,342 high quality (minor allele frequency > 2%) single nucleotide polymorphisms for genomic prediction. For grain yield, the experimental hybrids exhibited an average mid-parent heterosis of 40%. Genomic best linear unbiased prediction (GBLUP) for hybrid performance yielded an average prediction accuracy of 0.76-0.93 under the prediction scenario where both parental lines in validation sets were included in the training sets (T2). However, when only female tester was shared between training and validation sets (T1F), prediction accuracies declined by 12-90%, with plant height showing the greatest decline. Mean accuracies for predicting the general combining ability of male parents ranged from 0.33 to 0.62 for all traits. Our results showed hybrid performance for agronomic traits can be predicted with high accuracy, and optimizing genomic relationship is essential for optimal training population design for genomic selection in sorghum breeding.

Genetics of fertility restoration in A1 Cytoplasmic Genetic Male Sterility (CGMS) systems in Sorghum (Sorghum bicolor L. Moench)

The cause of Cytoplasmic Genetic Male Sterility (CGMS) is specific nuclear and mitochondrial interactions. Almost all commercial sorghum hybrids were developed using the A1 cytoplasmic genetic male sterility system. Understanding the inheritance of fertility restoration in sorghum for A1 cytoplasm, for example, can improve the selection efficiency of restorer lines for increased seed production. In a cross of male sterile line 296A with A1 cytoplasm and restorer lines comprised of a set of Recombinant Inbred Lines (RILs), the inheritance pattern of fertility restoration of sorghum was studied. The F1 hybrid was completely fertile, revealing the dominant nature of fertility restoration, which is controlled by one or two major genes with modifiers. In this study, the genetics of fertility restoration of the A1 cytoplasmic nuclear male sterility system (CGMS) in sorghum were investigated in segregating F2 and BC1 populations of A1 cytoplasm crosses. Fertility restoration was governed by a monogenic inheritance (3F:1S) mechanism represented by a single dominant gene responsible for fertility restoration in all of the crosses studied.

DNA marker analysis of flowering time, semi‐dwarf, fertility restorer, and brown midrib genes in sorghum (Sorghum bicolor [L.] Moench)

AbstractDNA markers for the genes for flowering time (Ma1/SbPRR37, Ma2/SbEHD1, Ma3/SbPhyB, Ma4/SbCO, Ma5/SbPhyC, and Ma6/SbGHD7), semi‐dwarfism (dw1 and dw2), fertility restorers (Rf1, Rf2, Rf5, and partial A1 mitochondrial sequence) and cytoplasmic–nuclear male sterile (CMS), and brown midrib (bmr‐2, bmr‐6, and bmr‐18) were developed in this study. These markers and previously published DNA markers for dw3 were used to analyze sorghum cultivars used in breeding programs in Japan and traditional landraces from the National Institute of Agricultural Sciences (NIAS) sorghum core collection. Agronomically useful mutations were effectively detected in the cultivars by the DNA markers. Most of the mutation types for flowering and semi‐dwarfism were detected in sorghum cultivars, but only the dominant‐type allele of SbEHD1 and SbPhyB for flowering was detected in the NIAS collection. Fertility restorers and CMS were rarely detected in NIAS collection. Agronomically useful traits such as early flowering and semi‐dwarfism became combined in the modern cultivars by phenotypic evaluation for the adaptation of harvesting to the temperate zones. Fertility restorers and CMS were introduced for F1 hybrid breeding. These phenotypes are often recessive, so information on the alleles responsible would be helpful for sorghum breeding. We expect these DNA markers to facilitate and improve the efficiency of F1 sorghum breeding programs.

Genetics of fertility restoration of the A5 cytoplasm in pearl millet (Pennisetum glaucum)

For the efficient hybrid seed production, several cytoplasmic-nuclear male sterility (CMS) systems were identified in pearl millet (Pennisetum glaucum ( L) R. Br). However, most of the hybrids developed in pearl millet are based on A1 cytoplasm due to availability of high frequency of restorers in the natural population. This dependence on a single cytoplasm makes any hybrid seed industry vulnerable to disease and insect pest epidemics. A5 can be a good alternative to A1 since it is the most stable cytoplasm available. But due to very low frequency (~2%) of restorers available in germplasm, utilization of A5 cytoplasm in hybrid development had been almost negligible. Restorer breeding efficiency of this CMS system can be significantly increased by understanding the inheritance of fertility restoration by Rf-gene for A5 cytoplasm, which is poorly studied so far and only one publication is available. Present investigation aims to understand the genetics associated with fertility restoration of the A5 system of CMS in pearl millet using pollen fertility and seed set percent as a criterion in determining the fertile and sterile plants. Three diverse cytoplasmic-nuclear male-sterile A-lines belonging to the A5 cytoplasm (ICMA502555, ICMA507999 and ICMA512222) were crossed with two restorer lines (A5RT-17/8 and A5 RT-17/26) to produce 6 F1s and their respective F2s and backcrosses. These were evaluated for male sterility (S) and fertility (F) at ICAR-IARI, New Delhi during kharif 2019 and segregation ratio was established in all the generations using χ2 test. The segregation pattern of malesterile (S) and male fertile (F) plants observed in F2 and BC1 indicated dominant single-gene control of male fertility restoration with ICMA502555 and ICMA507999. However, segregation pattern with male-sterile line ICMA512222 had a good fit to 15F:1S ratio indicating two gene control with duplicate interaction. The deviations from the expected ratio with ICMA512222 cross may be due to influence of modifiers. Association between pollen fertility and seed set percent was significant and positive.

The drivers and methodologies for exploiting wild Cajanus genome in pigeonpea breeding

With the exception of Cajanus cajan (L.) Millspaugh (pigeonpea), the remaining species of genus Cajanus have not been domesticated. For millennia these taxa have persisted in natural habitats through self-sown seeds. These wild species are an asset for sustaining future pigeonpea breeding programmes since they contain certain traits (genes) that are necessary for encountering various breeding challenges related to crop improvement and adaptation. In this review we identify the key traits from wild Cajanus species, and discuss various physical and genetic constraints encountered in their utilization in introgression breeding. Some noteworthy achievements recorded from inter-specific breeding programmes in pigeonpea are also discussed. These include the development of (1) high protein (> 28%) genotypes (2) cytoplasmic nuclear male sterility systems (3) highly (> 95%) self-pollinating genotypes, and (4) resistance sources to sterility mosaic disease, nematodes, salinity, photo-insensitivity, pod borers, podfly, bruchids, and Phytophthora blight. To help pigeonpea breeders engaged in inter-specific breeding programmes, we suggest the division of the secondary gene pool germplasm into two sub-group/tiers on the basis of ease in hybridization.

Maternal inheritance of male sterility in the progeny of a natural hybrid between Cajanus lineatus and C. cajan

AbstractAdoption of pigeonpea hybrids in central and southern India is showing high impact with on‐farm yield advantages of >30%. The hybrid pigeonpea technology, the first in any legume crop, is based on a cytoplasmic‐nuclear male‐sterility (CMS) system. For a long‐term sustainability of hybrid programme, it is imperative that both nuclear diversity and cytoplasmic diversity are maintained among hybrid parents. In this context, a continuous search for new CMS‐inducing cytoplasms is necessary. This paper reports detection of maternal inheritance of male sterility in the progeny derived from a natural hybrid between a wild relative [Cajanus lineatus (W. & A.) Maesen comb. nov.] of pigeonpea and an unknown pigeonpea [Cajanus cajan (L.) Millsp.] genotype. In the present study, the male sterility was maintained up to BC7F1 generation by an advanced breeding pigeonpea line ICPL 99044. This male sterility inducing cytoplasm of C. lineatus was tagged as A6. In future, this CMS genetic stock can be used to develop a range of new pigeonpea hybrids with high yield and adaptation.

Inheritance of fertility restoration of A5 cytoplasmic-nuclear male sterility system in pearl millet [Pennisetum glaucum (L.) R. Br.

The A5 cytoplasmic-nuclear male sterility (CMS), the most stable cytoplasm available in pearl millet [Pennisetum glaucum (L). R. Br.] is yet to be utilized on large scale by breeders owing to limited fertility restoration in the available germplasm for this cytoplasm. An understanding of the inheritance of fertility restoration can make significant contributions to restorer breeding efficiency of this CMS system. This study investigated inheritance of fertility restoration of this CMS system in which three diverse isonuclear A-lines (P1) were crossed with two A5 restorer lines (P2) to produce 4 F1s and their respective F2s and two backcrosses. These were evaluated for male sterility (S) and fertility (F) for two seasons at International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) Patancheru. The segregation patterns in the F2 (54F:10S) and BCP1 (3F:1S) were broadly suggestive of trigenic inheritance of male fertility restoration, where dominant alleles at any two of the three duplicate complimentary loci will lead to fertility restoration.

Key biological factors related to outcrossing-productivity of cytoplasmic-nuclear male-sterile lines in soybean [Glycine max (L.) Merr.

Soybean scientists in China have tried to utilize heterosis for yield enhancement. A number of cytoplasmic-nuclear male-sterile (CMS) lines, their maintainers and restorers were developed, but the commercialized hybrid seed production is still a bottleneck. Five soybean CMS lines with their maintainers and restorers were studied for the sufficiency of viable and effective pollens from maintainers/restorers, the natural insect pollinators in fields and outcrossing capacity of CMS lines. The results showed that the maintainers/restorers may offer viable pollens during the flowering day time in Nanjing and Taiyuan (germination rate 77.8–86.5%). But the morning dew causes pollens losing effectiveness in Nanjing, while normal in Taiyuan, indicating the location and planting season without morning dew is a key for CMS/hybrid seed production. In CMS fields in Taiyuan, the major insect pollinators composed of China bee (54.3%), Mellifera bee (11.3%), Northern-cutting bee (6.0%), Corn belt hair bee (1.5%), etc. A parallel trend between insect number and flower nectar amount in a day with peak time at 12:00–13:00 implied the soybean nectar attracting the insects. The outcrossing pod-set rate of CMS lines in net-room with artificial-raised bees, net-room without bees and open-field with enhanced natural insects was 57.8, 2.1 and 47.3%, respectively, indicating that insect pollination is necessary. The realized outcrossing pod-set rate in seed production field achieved 88.9% (1460.4 kg/ha) for SXJLCMS1A × SXJLCMS1B and 145.2% (1829.5 kg/ha) for SXCMS1A × JY-31, different outcrossing capacity existed among CMS lines. Thus the CMS/hybrid seed production can be improved through optimizing the pollen effectiveness, pollinator population and talented CMS line.

Retrospective and perspective on <italic>indica</italic> three-line hybrid rice breeding research in China

China’s first indica hybrid rice technology has opened up new methods to substantially increase rice yield, tremendously contributing to China’s food security. In this article, we review the discovery of wild abortive male sterility in rice and the history of the development and combination of the male sterile line, maintainer line and restorer line (the “three-line”). With regards to China’s large-scale application of cytoplasmic-nuclear male-sterile (CMS) lines, in addition to the wild-abortive CMS line, there are also Gang, D, Yinshui, dwarf-abortive, Honglian and K types, of which, only the Honglian type exhibits a different restoring and maintaining relationship from that of the wild-abortive line. The diversity of cytoplasmic sterile lines of hybrid rice solves the potential risk in production caused by the use of a single cytoplasmic line. With respect to the breeding of elite parents, the breeding and application of fine male sterile lines, such as Zhenshan 97A, V20A, II-32A and Jin 23A, and fine restorer lines, such as IR24, Minghui 63 and Shuhui 527, were introduced. Currently, the major varieties of China’s three-line hybrid rice have transformed from the high-yielding varieties in the past into the current high-quality, high-yield and disease-resistant varieties. However, three-line hybrid rice faces several problems, such as slow increases of yield potential, low efficiency in genetic improvement of maintainers, the high cost of hybrid rice seed production and the lack of adaptability to the simplified cultivation method. To meet these challenges, it is recommended to establish objectives of breeding new hybrid rice varieties with high quality, high yield, high safety and high efficiency by applying molecular tools to hybrid rice breeding and developing “three-line” hybrid rice parents suitable for mechanized seed production. These would significantly enhance the technological competitiveness of hybrid rice.

From concept to field: evolution of hybrid pigeonpea technology in India

The role of hybrid vigour in the enhancement of productivity is well recognized. Its application, however, has been restricted to a few crops and legume breeders could never take its advantage due to various seed production issues. Recently, plant breeders have succeeded in creating the world's first commercial hybrid in a food legume, popularly known as red gram or pigeonpea [Cajanus cajan (L.) Millsp.]. This was possible due to success in breeding a stable cytoplasmic nuclear male sterility (CMS) system and exploitation of its partial natural out-crossing. This development has provided golden opportunities to break the decades-old yield plateau in this crop. Among the hybrid combinations evaluated, ICPH 2671 and GTH 1 were found most promising. In the on-farm trials ICPH 2671 recorded 46.5% superiority over the best available cultivar Maruti; while GTH 1 was 33.9% superior to the control, and it was released in Gujarat state in 2004; but for some reasons associated with its stability of fertility restoration it failed to reach the level of commercialization. The outstanding performance of hybrid ICPH 2671 led to its release by both, a private seed company as ‘Pushkal’ and by the state Government of Madhya Pradesh as ‘RV ICPH 2671’ in 2010. The journey of the evolution of hybrid pigeonpea technology was fascinating and challenging. It took a long time of over 39-years from its conceptualization to finally reaching Indian farmers. The impact of this breakthrough was visible when two more hybrids ICPH 2740 and ICPH 3762 were recently released in India. Since it is a path breaking research among food legumes, an attempt has been made here to archive the milestones of this historic plant breeding event.

Marker-assisted breeding for rf1, a nuclear gene controlling A1 CMS in sorghum (Sorghum bicolor L. Moench)

A study on marker-assisted selection (MAS) for the rf1 gene, which controls pollen sterility in the sorghum A1 cytoplasm, was conducted on the offspring population of two crosses between a maintainer line, BTx-622, and two sweet sorghum lines, BJ-299 and Lunen-2, to test the effectiveness of the MAS method and develop maintainer lines with sweet and juicy stalks and corresponding cytoplasmic-nuclear male sterility (CMS) lines. The simple sequence repeat marker Xtxp18 exhibited a high accuracy (95.098 %) for selecting recessive homozygotes for the rf1 gene. The segregation ratio matched the expected ratio calculated according to the reported genetic distance in the F2 population of the two crosses used. Finally, four excellent maintainer lines/CMS line pairs (F5/BC3) with high stalk juice and stalk juice sugar contents were developed. The MAS method based on Xtxp18 for the sorghum rf1 gene could be used for hybrid breeding programs at a low cost in the future.

ICPH 2671 – the world's first commercial food legume hybrid

AbstractICRISAT scientists, working with Indian programme counterparts, developed the world's first cytoplasmic‐nuclear male sterility (CMS)‐based commercial hybrid in a food legume, the pigeonpea [Cajanus cajan (L.) Millsp.]. The CMS, in combination with natural outcrossing of the crop, was used to develop viable hybrid breeding technology. Hybrid ICPH 2671 recorded 47% superiority for grain yield over the control variety ‘Maruti’ in multilocation on‐station testing for 4 years. In the on‐farm trials conducted in five Indian states, mean yield of this hybrid (1396 kg/ha) was 46.5% greater than that of the popular cv. ‘Maruti’ (953 kg/ha). Hybrid ICPH 2671 also exhibited high levels of resistance to Fusarium wilt and sterility mosaic diseases. The outstanding performance of this hybrid has led to its release for cultivation in India by both a private seed company (as ‘Pushkal’) and a public sector university (as ‘RV ICPH 2671’). Recent developments in hybrid breeding technology and high yield advantages realized in farmers' fields have given hope for a breakthrough in pigeonpea productivity.

Genetics of fertility restoration of the A<sub>4</sub> cytoplasmic-nuclear male sterility system in pearl millet

Inheritance of fertility restoration of the A<sub>4</sub> system of cytoplasmic-nuclear male sterility in pearl millet was investigated using six crosses between two diverse male sterile lines (A-lines) and three diverse restorers (R-lines). The segregation pattern of male sterile (S) and male fertile (F) plants observed in F<sub>2</sub>, and BC<sub>1 </sub>in two seasons at ICRISAT, Patancheru, indicated the dominant single-gene control of male fertility restoration. The segregation pattern in BC<sub>1</sub>F<sub>2</sub> progenies derived from the fertile BC<sub>1</sub> plants evaluated for one season provided further evidence for the single-gene control. The season did not have much effect on fertility restoration. The information on the single-gene control of fertility restoration will help in diversifying the restorer genetic base of the A<sub>4 </sub>CMS system and enhance R-line breeding efficiency in pearl millet.

Evidence of a unique inter-allelic epistatic interaction for seed coat color in pigeonpea [Cajanus cajan (L.) Millspaugh

For enhancing productivity and profitability of pigeonpea (Cajanus cajan (L.) Millsp.) a hybrid breeding technology based on a cytoplasmic nuclear male-sterility (CMS) system was developed at International Crops Research Institute for the Semi-Arid Tropics (ICRISAT). Among the elite hybrids ‘ICPH 2671’ was most out-standing with 30–35% yield advantage over local varieties. Despite its commercial release in 2010, this hybrid could not attract many growers due to its less preferred seed color. Both the parents (ICPA 2043 and ICPR 2671) of this hybrid and their crossed seeds had Greyed-orange [code no*: 167B/C (* The standard identification numbers of Royal Horticultural Society)] color. However, the commercial seeds produced on the hybrid plants surprisingly have Greyed-purple (code no: 183C) color that fetches relatively less profits. To understand the genetics of this phenomenon a cross combination was studied for seed color in F1, F2, and B1C1F1 generations. The data suggested that three independent dominant genes controlled the expression of seed coat color. The observations further suggested the presence of two dominant genes (Brsd1 and Brsd2) in the male-sterile line ‘ICPA 2043’, ‘ICPR 2671’ and another dominant gene (Brsd3) in the restorer line. In the hybrid plants genes Brsd1 and Brsd2 act in complementation with the third gene Brsd3, this also acts as a modifier. The presence of Brsd3 gene in a genotype together with Brsd1 and/or Brsd2 produces a strong inter-allelic epistatic interactions resulting in the Greyed-purple (183C) seed color. It was further postulated that the genetic constitution of ‘ICPA 2043’ is Brsd1Brsd1Brsd2Brsd2brsd3brsd3, while the genotype of the restorer line ‘ICPR 2671’ is brsd1brsd1brsd2brsd2Brsd3Brsd3.

Ensuring Genetic Purity of Pigeonpea Hybrids by Incorporating a Naked‐Eye Polymorphic Marker in A and B Lines

ABSTRACTTo enhance the productivity of pigeonpea [Cajanus cajan (L.) Millspaugh] a hybrid breeding technology, based on the cytoplasmic nuclear male‐sterility (CMS) system and partial natural outcrossing, is currently been used. However, there are difficulties to maintain genetic purity of the hybrids and their parents. The incorporation of an easily identifiable morphological marker (naked eye polymorphism [NEP]) could be used to determine seed purity. The morphological marker selected for this study, obcordate leaf, is not present in cultivated pigeonpea; it is inherited as a single recessive gene and can be observed soon after planting (approx. 6 wk). To incorporate the obcordate leaf shape into hybrid parents, the trait was transferred from the germplasm accession ICP 5529 into male‐sterile (A lines) and the corresponding maintainers (B lines). The hybrids derived from crosses involving obcordate leaf A lines and normal leaf fertility restorers (R lines) were fully fertile and had normal lanceolate leaves; thus the difference between A line and hybrids was clear. The use of obcordate leaf as a NEP marker in pigeonpea would contribute to preserve parental line purity and confirm hybrid status.

Genetics of Fertility Restoration in A4‐Based, Diverse Maturing Hybrids of Pigeonpea [Cajanus cajan (L.) Millsp.

ABSTRACTRecently, a breeding technology for hybrid pigeonpea [Cajanus cajan (L.) Millsp.] based on cytoplasmic‐nuclear male sterility (CMS) and partial natural outcrossing was developed at ICRISAT. The limited number of experimental hybrids tested has shown the presence of a considerable amount of hybrid vigor for seed yield. In the present study, one extra‐early‐ (120 d), two early‐ (150 d), and two late‐maturing (180 d) pigeonpea hybrids were studied to generate information on the genetics of fertility restoration of the A4 CMS system. In the extra‐early‐maturing hybrids, pollen fertility was controlled by a single dominant gene, whereas in the early‐ and late‐ maturing hybrids, male fertility was governed by two duplicate dominant genes. It was also observed that hybrids with two dominant genes produced a greater pollen load and expressed greater stability as compared with those carrying a single dominant gene. The information on the inheritance of fertility restoration will help in designing strategies for breeding elite hybrid parents, and it was concluded that for breeding hybrids with stable fertility restoration, the presence of two dominant genes is essential.

Mapping Male Fertility Gene with SSR Markers in Parents of Cytoplasmic- Nuclear Male-Sterile Line NJCMS3A in Soybean

Utilization of the three lines [cytoplasmic-nuclear male-sterile(CMS) line,maintainer line,and restorer line] in producing hybrids with heterosis has been very successful in a number of crops including soybean.The soybean CMS line NJCMS3A was developed through six consecutive backcross generations with N21566 as cytoplasm source and N21249 as nuclear donor.The objective of our study was to reveal the genetic mechanism and mapping the male fertility gene in the two parents of the CMS line NJCMS3A,i.e.N21566 and N21249.The male fertility of(N21566×N21249) F_2 and BC_1F_1 showed that the segregation ratio of fertile to sterile plants fitted the expected ratios of 3:1 and 1:1,respectively,indicating one pair of gene conferring male fertility in the parents of NJCMS3A with dominance allele in N21249 being male fertile.This gene might be one of the restorer genes for the male sterility of NJCMS3A.The mapping results on BC_1F_1 and F_2 showed that out of the 793 randomly selected SSR markers,Satt331,CSSR133 and Satt477 on linkage group O were linked to the male fertility gene Rf with their genetic distances of 8.1～10.4,11.4～16.4,and 13.3～19.2 cM,respectively,in a same order and its location between Satt331 and Satt477.

Application of SSR markers for molecular characterization of hybrid parents and purity assessment of ICPH 2438 hybrid of pigeonpea [Cajanus cajan (L.) Millspaugh

With an objective of achieving a breakthrough in the productivity of pigeonpea, a hybrid breeding technology based on elements of the cytoplasmic-nuclear male-sterility (CMS) system and partial natural out-crossing has recently been developed. However, there is no molecular diversity information available on parental lines of hybrids being generated at the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT). This study deals with the use of 148 simple sequence repeat (SSR) markers, including 32 novel markers reported here for the first time, on 159 A (cytoplasmic male sterile), B (maintainer) and R (fertility restorer) lines. In total, 41 (27.7%) markers showed polymorphism with 2 to 6 (average 2.6) alleles and 0.01 to 0.81 (average 0.34) polymorphism information content (PIC) value. Of these polymorphic markers, 22 SSR markers showed polymorphism between A (ICPA 2039) and R (ICPR 2438) lines of the commercial hybrid (ICPH 2438); however, only 21 of these SSR markers showed the same profile between A (ICPA 2039) and B (ICPB 2039) lines. Finally, two SSR markers, CCB4 and CCttc006, were found most suitable for purity assessment of hybrid seeds of the ICPH 2438 hybrid. The utility of these two diagnostic SSR markers has been demonstrated by using seed lots of this hybrid from two sources, ICRISAT and Mahabeej. It is anticipated that molecular diversity information generated on parental lines of hybrids under development, and identification of the two most suitable markers for testing the purity of hybrid seeds of ICPH 2438, will facilitate the pigeonpea hybrid breeding programme.