Construction Of Rice Mutant Library And Preliminary Study On OsARP Gene Function

Rice is one of the most important food crops in China and plays a decisive role in ensuring national food security.With the completion of rice whole-genome sequencing,the discovery of new genes and understanding of the functions of genes has become a top priority for functional genomics research,and the creation of excellent rice germplasm resources has laid a solid foundation for rice functional genomic research and rice breeding.In this experiment,the seeds of rice Nipponbare were used as materials.The mutants were obtained by using the chemical mutagen ethyl sulfonate(EMS)at different concentrations and different times for seed soaking and phenotypic screening of the M2 and M3 generation populations.Expression,gene knockout and gene editing techniques were carried out to study the gene function of rice Os ARP(Oryza sativa Auxin responsive protein),and the test results showed that:1.Treat rice with 7 EMS mutagenesis concentrations(0.25%,0.5%,0.75%,1%,1.25%,1.5%)and 4 mutagenesis times(6 h,12 h,18 h,24 h)According to the statistics of seed germination rate,it was found that with the increase of EMS mutagenesis concentration and mutagenesis time,the germination rate of rice seeds gradually decreased,indicating that the EMS reagent had a delay and inhibitory effect on seed germination,and even lethal effect;by analyzing the mutagenesis concentration With the curve of mutagenesis time on the germination rate of rice seeds,the optimum dosage of mutagenesis treatment was finally determined: 0.5% EMS concentration and mutagenesis time of 12 h.2.Through EMS mutagenesis treatment,8 variant types of 355 M1 generation single plants were obtained,including tiller number,plant height,leaf color,leaf shape,growth period,seedless fruit,grain size,ear tip degradation,etc.After phenotypic genetic analysis of M2 and M3 mutants,a total of 248 agronomic traits including 8 tillers,plant height,leaf color,leaf shape,growth period and spike shape were observed in some agronomic traits of rice plants.The mutation frequency is9.6%.3.To verify the function of this gene by transferring the overexpression vector and gene editing vector constructing Os ARP gene into the Arabidopsis genome using the Agrobacterium dipping method.The results showed that: 10 Arabidopsis transgenic plants with overexpression vector p CAMBIA1305-Ubi-ARP were obtained,6 of which amplified target gene fragments by PCR;10 Arabidopsis transgenic plants with gene editing vector p HSN401-Os ARP were obtained,10 The strain can amplify the hygromycin fragment by PCR;by observing the phenotype,it is found that the height of the transgenic plant is significantly lower than that of the wild type,which is stunted,and the transgenic plant has smaller leaves and more pods,and the gene is found.After the editing vector was transformed into Arabidopsis thaliana,the leaf color was lighter to light green.