Inheritance of fertility restoration in Indian onion (Allium cepa L.)
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Date
2024
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Punjab Agricultural University
Abstract
Onion is a cross-pollinated crop and is suitable for heterosis breeding to develop elite F1 hybrids. The small size of onion flowers hinders the use of manual emasculation and pollination techniques but could be alleviated by adopting the mechanism of cytoplasmic male sterility. The development of A (Smsms) and B (Nmsms) lines is time consuming and laborious process through conventional approaches but can be accelerated through marker-assisted selection. The existing molecular markers can accurately predict cytotypes, but have been found inadequate in predicting the Ms locus in Indian populations. This indicated the possibility of another fertility restoration locus (Ms) in Indian onion populations. To explore this, the study titled "Inheritance of fertility restoration in Indian onion (Allium cepa L.) was conducted for two consecutive years- Rabi 2021 and Rabi 2022. The parental lines namely, male sterile (D-121-A), maintainer (D-121-B) and restorer (R-230) lines were characterized through morphological, microscopic, and molecular analyses. The visual assessment indicated that male sterile plants exhibited shrunken or distorted anthers in A-line, while male fertile plants displayed plumpy anthers in B and R-lines. Subsequent microscopic analysis using acetocarmine validated these observations, as the sterile plants showed no staining, whereas the anthers of male fertile plants exhibited a distinct red coloration. To conduct segregation analysis, a segregating F2 progeny of the cross-involving CMS A-line (D-121-A) and the R-line (R-230) was used. Phenotyping and chi-square tests conducted on the F2 population indicated a deviation from the expected Mendelian ratio of male fertile to male sterile (3:1). The observed ratio demonstrated a good fit in a 9:7 ratio, suggesting the presence of two dominant complementary genes regulating the fertility restoration locus (Ms) in Indian population. Cytotype markers based on gel electrophoresis (accD and MKFR) and the nuclear-specific marker (AcPMS1) demonstrated accuracy in predicting the N/S cytotypes and Ms locus in the parental lines. However, when genotyping was done in F2 population with AcPMS1, it was observed that out of 160 individuals, 22 plants did not accurately predict the Ms locus. The genotypes of these 22 individuals did not match with their phenotypic appearance. This discrepancy suggested the potential existence of a recombination event between the marker and the QTL (Ms locus). Furthermore, gel-based markers were found to be time-consuming and had low throughput. Consequently, a high-throughput KASP assay was devised to identify cytotypes and the fertility restoration locus (Ms). The KASP based cytotype markers (OC_17 and OC_19) and SNP based nuclear marker (OMs_1 and OMs_2) were utilized to distinguished the cytotypes and Ms locus in parental lines, respectively. OC_17 and OC_19 accurately predicted the cytotypes of A, B and R parental lines and showed complete resemblance with gel-based markers. However, the SNP-based markers used in the F2 population showcased varying results. OMs_1 displayed clear and distinct cluster formation, while OMs_2 showed distorted polymorphism in the F2 population. The accuracy percentages were 84.82% for OMs_1 in predicting the nuclear Ms locus. The developed KASP based marker (OMs_1) has comparable marker efficiency with the gel-based marker (AcPMS1). As Pran a triploid Allium species is considered as a source of S cytoplasm based on restriction assays in this investigation both gel and KASP-based cytotype markers identified 'Pran', along with A and R-lines, as S-cytoplasm. This provides further support for the idea that the sterile cytoplasm in Indian onion populations possibly introduced from the "Pran". This study being the first report regarding fertility restoration control in Indian onion further emphasizes on mapping genomic regions governing fertility restoration for generating tightly linked markers to enhance marker-assisted breeding programs in Indian onions.
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Kavita (2024). Inheritance of fertility restoration in Indian onion (Allium cepa L.) (Unpublished M.Sc. thesis). Punjab Agricultural University, Ludhiana, Punjab, India.