Functional Analysis And Breeding Potential Of Rice F-box Gene OsFBX76

Rice is one of the most important food crops and increasing the yield has been the main problem in our country.With the breakthrough of 'dwarf breeding' and 'hybrid technology',the yield rose to a higher level.However,the yield encountered a bottleneck in the last decade.The progress of urbanization and conflicts between people and food need higher demands on rice yield.The 'molecular design breeding'strategy will provide an efficient approach for improvement of rice yield and the discovery of genes associated with yield traits is the foundation of high yield molecular breeding.F-box proteins are components of E3 ubiquitin ligase S-PHASE KINASE-ASSOCIATED PROTEIN,CULLIN,F-BOX CONTAINING COMPLEXES(SCF),which are widely founded in eukaryotes.The F-box proteins mediate a wide variety of biological processes such as plant stress response,hormone signaling,self-incompatibility,optical signal transduction and floral organ development.In this study,we characterized a F-box gene OsFBX76(LOC_Os03g02550)in rice.Several homozygous and stable RNAi and overexpression transgenic lines under NIP background were generated and used for functional analysis.The molecular mechanism of OsFBX76 for regulating rice grain yield was preliminarily revealed and the main results are as follows:1.Yeast two-hybrid experiments showed that Skp1 interacts with OsFBX76 protein.2.Using quantitative real time PCR(qRT-PCR),the expression pattern of OsFBX76 was analyzed.The results showed that OsFBX76 constitutively expressed in all tissues,including leaf,leaf sheath,internode,stem and young panicle.The highest transcripts were detected in leaves,and the expression level of OsFBX76 declined with the development of panicles.3.At maturity,several yield traits were measured.1000-grain weight,the yield per plant and seed setting rate decreased by 8.15%?19.70%,48.57%?70.65%,and 27.37%?60.03%,respectively,in overexpression lines.By contrast,1000-grain weight,the yield per plant and seed setting rate increased by 4.93%?7.67%,7.84%?17.81%and 4.62%?7.27%,respectively,with the decreased expression levels of OsFBX76 in RNAi lines.Therefore,OsFBX76 negatively regulate seed size and seed setting rate to affect rice yield.4.Histomorphological analysis indicated that OsFBX76 gene negatively regulates cell elongation rather than cell division and resulted in a shorter second internode from top than wild type.Scanning electronic microscopy(SEM)analysis of spikelet hull suggested that OsFBX76 negatively modulate cell length.Therefore,OsFBX76 negatively regulate cell size.5.Using CRISPR/Cas9 system,homozygous mutants of OsFBX76 were obtained under the background of Wuyunjing 7(WYJ7)and suken 118(SK118),which provide useful materials for further researches on functional analysis and breeding potential.