Map-based Cloning Of MULTI-FLORET SPIKELET 2(MFS2) Gene In Rice(Oryza Sativa L.)

Rice(Oryza sativa L.) is one of the most important food crops worldwide and a model monocot plant for gene function analysis. It is of great significance to study its flower development. Researches help us understand the forming mechanism of rice spikelet or flower in reproductive development process. Meanwhile, it also provides theory basis for inflorescence research of other flowering plants. So, it is a good way to improve rice yields and enrich knowledge of plant growth and development biology.In the present study, we reported named a mutant multi-floret spikelet 2(MFS2) in rice.We investigated the mfs2 mutants with phenotype, histological and cytological observation, did map-based cloning, expression pattern analysis and functional verification of MFS2 gene. The results were as follows:1. Phenotypic observation of mfs2 mutants At flowering stage, the mfs2 mutants showed palealess or curving-palea or lemma-like palea; extra elongated glume-like organ; partial ectopic degenerated stamens or increased stamens; partial carpels ectopic, and three stigmas in very few occasions.2. Molecule mapping of MFS2 In this study, the mutant trait was controlled by a single recessive gene. MFS2 was mapped between two simple sequence repeat markers RM17349 and RM17391 on 4th chromosome, and the genetic distances were 0.001 c M and 0.020 c M respectively, physical distance was about 1.02 Mb.3. Cloning and protein analysis of MFS2 DNA and c DNA sequencing results showed that gene Os04g47890, which encoded a MYB transcription factor, had a C-base deletion in the 5th exon of mfs2, causing the 213 th amino acid into the termination codon. Therefore, we identified Os04g47890 as the candidate gene.4. Function analysis of the MFS2 gene The MFS2 RNAi vector was established and transformed into Zhonghua11. We detected the transgenic positive plants by PCR and q PCR, and found the expression level of MFS2 was decreased. Spikelets of those transgenic plants showed similar phenotypes as mfs2. The results demonstrate that the Os04g47890 is the MFS2 gene.5. Expression pattern analysis of the MFS2 gene, phylogenetic analysis and protein subcellular localization q PCR was used in expression pattern analysis. The results showed that MFS2 gene was expressed at different stages in various tissues including root, stem, leaf, sheath, panicle and floral organs, especially high in early heading stage. Clustering analysis indicated that MFS2 belonged to a clade of MYB-1R(MYB-related gene) in the MYB family, and was located in the nucleus.6. The expression analysis of relative floral development genes We analyzed the expression levels of relative floral development genes between wild-type and mfs2 by q PCR. Those results displayed that the expression level of the floral development gene FZP was up-regulated, while the expression of others were varied in different stages. In addition, the expression level of the floral organ development genes was also in the same case. But overall, those genes’ expression was consistent with phenotype of mfs2.