Phenotypic Characterization And Fine Mapping Of A Male Sterile Mutant, XS1, In Rice (Oryza Sativa L.)

XS1 was derived from a spontaneous mutation and confirmed to be a no-pollen type mutant of male sterility in rice.The floret of the mutant,consisting of six stamens and one pistil,looks the same as that of the wild type in the male-female organs,except that the filaments are long and thin,and the anthers are withered in white transparence.It is confirmed that XS1 is a none-pollen-type mutant of male sterility for no pollen grains can be stained with I₂-KI solution and the anther locules are always hollow.Anther transverse sections indicate that the mutant microspores are abnormally condensed and agglomerated to form a deep-stained cluster at the late microspore stage,which results in the ceasing of the vacuolation process of microspores,and,therefore,the mutant forms no functional pollens for reproduce.During the heading stage,all the individual plants in the F₁ and F₂ progenies from the crosses between XS1 and other normal rice lines were investigated.In the four F₁ progenies,all plants exhibited wild-type phenotype,suggesting that the mutant trait is recessive.In the four F₂ populations,all the segregation rates of fertility and sterility plants fit the ratio of 3:1,and,additionally,in the 3 BC₁F₁ populations,all the segregation rates of fertility and sterility plants fit the ratio of 1:1.Genetic analysis processed in 4 F₂ populations and 3 BC₁F₁ populations reveal that the mutation is controlled by a single recessive gene,and it is termed as vrl(Vacuolation retardation l).The polymorphisms between XS1 and other rice lines G630 and M63 were examined with 512 SSR markers and the most polymorphism richment population XS1/G630 was selected for mapping.Firstly,total 103 SSR polymorphic markers were selected and used to survey in a small populations which was composed of the two parents,four of wild type F₂ plants,and six of F₂ mutants plants.The result showed that 3 SSR markers RM470,RM303 and RM317 located on chromosome 4 were obviously associated with the XS1 phenotype.Then,the three markers were used to survey all the mutant plants in the same F₂ population,and were all verified to be linked to vrl with the genetic distance of 3.4,2.6 and 2.4 cM,respectively.In order to fine-map the vrl gene,two large segregation populations were generated from crosses of XS1/G630 and XS1/M63,respectively.Besides,sevaral SSRs,InDels markers were newly developed according to the publicly available rice genomic sequences.Reconbination analysis indicated that the vrl gene was finally located within a genetic interval of 0.27 cM,flanked by markers FID30,FS15,and co-segregated with marker FC 4-2.The delimitation region,according to the japonica rice genomic data,was estimated to be a 48 kb physical interval.Within the 48 kb physical interval,totally 8 putative genes were predicted by some sofiwares of the bioinformatics.LOC_Os04g51130 is a expressed protein with unknown function;LOC_Os04g51080 is a scramblase protein;LOC_Os04g51090 is a tRNA-splicting endonuclease positive effector-related protein;LOC_Os04g51100 is a putative COBW domain containing protein;LOC_Os04g51110 is a putative WD repeat-containing protein;LOC_Os04g51120 is an ENTH domain containing protein; LOC_Os04g51140 is an E2F-related protein;and LOC_Os04g51150 is a putative unclassified transposon protein.