关键词: 水稻, 直立短穗突变体, 基因克隆, 进化分析, 转录组分析

Abstract:

【Objective】In this study, we aimed to identify the ESP gene, whose mutation caused a phenotype, namely erect and short panicle, and to determine its regulatory role in the gene network that controls the related agronomic traits (e.g., plant types and panicle length).【Method】In this study, agronomic traits, such as plant height, panicle length and grain length at mature stages, were used as phenotypic marks to trace the esp mutant. Individuals carrying mutant phenotypes were selected from the F₂ population to cross with indica and japonica for further gene mapping and genome sequencing that were used to map the potential mutation region/sites. The bioinformatics software was used to analyze phylogenetic tree and gene expression. The total RNAs isolated from wild type and mutants were used for transcriptome RNA-seq analysis. The differential expressed genes and expression levels of genes related to plant hormone signal transduction and protein processing in endoplasmic reticulum were analyzed by GO software and KEGG database. The transcriptome data were verified by qRT-PCR.【Result】Phenotypic analysis showed that the esp mutant exhibited a erect panicle architecture. The plant height, panicle length, grain length and the number of spikelets per panicle were decreased in the esp mutants, when compared to wild type control, whereas grain width and the 1 000-grain weight were increased, although no obvious difference in the number of effective panicles between mutants and control. The ESP gene was mapped to a 7.58 Mb interval between markers C7-11 and C7-14 on the long arm of rice 7th chromosome by using the F₂ population of esp mutant and PA64. Genome sequencing demonstrated that a single nucleotide change (G to A) at the junction of 6th intron and 7th exon of LOC_Os07g42410, which led to a splicing defect, causing premature protein translation. The ESP was allelic to OsDEP2/OsEP2. Phylogenetic analysis revealed that the ESPs are widely present in monocot and dicot plants. Expression analysis predicted that ESP gene was highly expressed in stem, inflorescence, pistil, glumelle, lemma and ovary, and the expression level was gradually decreased with the ovary inflation. Transcriptome RNA-seq analysis of young panicle identified 630 differential expressed genes in esp mutants versus wild type, including 235 up-regulated and 395 down-regulated. GO and qRT-PCR analysis revealed that genes involved in plant hormone signal transduction and protein processing in endoplasmic reticulum were misregulated in esp mutants.【Conclusion】The ESP gene was allelic to OsDEP2/OsEP2, the mutation of which leads to multiple phenotypes, such as decreased plant height and shorter panicle length. Transcriptome analysis suggested that the ESP gene might affect plant development by regulating genes expression associated with plant hormone signal transduction and protein processing in endoplasmic reticulum.

Key words: rice (Oryza sativa L.), erect and short panicle mutant, gene cloning, phylogenetic analysis, transcriptome analysis