Identification and Candidate Gene Analysis of the ABNORMAL HULL 1(ah1)Mutant in Rice(Oryza sativa L.)
[Objective]Rice is the staple grain crop worldwide,and the morphology of its grains directly influences its ultimate yield,nutritional excellence,and economic significance.Moreover,the intricate interplay between floral development and grain morphology adds further significance to this relationship.Thus,exploring novel rice floral development regulatory genes and molecular regulatory mechanisms lays the foundation for larger and more plump grains rice varieties.[Method]Ethyl methyl sulfonate(EMS)was used to mutate XD1B(xian-type maintainer line),and a dwarf mutant abnormal hull 1(ah1)with abnormal formation of glume and lodicule was identified.The agronomic traits of both the wild-type and mutant were observed and recorded.Spikelets from various flowering stage were collected to histological and morphological analysis.The F2 segregating population was established by ah1 and 56S(xian-type thermo-sensitive sterility line),and utlized for genetic analysis and gene mapping.RNA was isolated from young panicles of both the wild-type and mutant,then reverse transcribed into cDNA.The RT-qPCR analysis was performed to analyze the relative expression levels of the genes regulating floral development and the key genes in the ABA synthesis pathway.[Result]The observation of agronomic traits revealed that the dwarfed plant was caused by the dramatic shortening of the internodes.At the same time,the mutant is also accompanied by severe spikelet abnormalities and low fruit setting rate.Histological and morphological analysis revealed that the ah1 mutant spikelets exhibited varying degrees of degeneration in floral organs such as palea,lemma,lodicules,and stamens.Some severely affected spikelets displayed altered floral organ characteristics and determinacy of floral meristems,often accompanied by extensive whitening.Based on the extent of degeneration,these spikelets could be classified as slight or severe mutant phenotypes.Genetic analysis showed a segregation ratio of 3﹕1 for the wild-type and mutant within the segregating population,indicating that the mutant traits of ah1 were controlled by a single recessive locus.The AH1 was mapped between the molecular markers RM6716 and RM128 on the chromosome 1,with a physical distance of approximately 8 Mb.Resequencing analysis of the mutant revealed that the LOC_Os01g53450 and LOC_Os01g51860 within this interval showed variation between wild-type and mutant,thus these two genes were provisionally identified as candidate genes.RT-qPCR analysis revealed significant alterations in the relative expression levels of floral organ development regulatory genes during the early developmental stages of mutant panicles;meanwhile,the relative expression levels of OsNCED1/OsNCED2/OsNCED3/OsNCED4/OsNCED5,the ABA synthesis pathway key genes,were severe inhibited.[Conclusion]AH1 plays a crucial role in the morphological formation of floral organs,such as palea and lemma in rice.LOC_Os01g53450 and LOC_Os01g51860 were provisionally identified as candidate genes in this work.
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