Study On Mutant And Application Of Rice By Space Mutation

A mutant of Indica rice H9808with orange-red trait was obtained by space mutation technology. The variation of the mutant was analyzed by AFLP molecular markers and genome re-sequencing technology. The gene controls orange-red trait of the mutant was fine mapped by SSR, InDel and CAPS markers. We used the mutant in rice breeding and new CMS lines with orange-red trait were obtained.The main results are as follows:1. AFLP molecular marker. analysis of mutant of space-flight in riceCompared with the wild-type material planted in the field, we found that the mutant dispalys young leaves with orange-red veins at the stage of panicle initiation its internode and one third of the down part stem become orange-red by the end of booting stage, and the color of the mutant hull is also orange-red from heading to maturity. The full grains of mutant reduced significantlyand become more narrower and longer, while other traits did not change significantly. Comparing the changes of mutant and wild-type material in genomic DNA by132AFLP molecular markers, we found52polymorphic fragments in the genome of the mutant, which showed that the mutant had changed both in the phenotype and the genome.2. Re-sequencing of the mutant and wild-typeTaking the indica rice9311as the basic sequence, we re-sequenced genome of mutant and wild-type material and found that the mutagenic effects of space mutation factors for the rice genome were basically distributed evenly and the mutation rate was positively correlated with chromosome size. The mutagenesis types are SNPs, InDel and SV. Their mutation rates are29.58%,7.49%,3.75%respectively. The percent of Base A in SNPs Mutant is24.4%, and the percent of A into T, C and G is12.30%,15.57%,72.13%respectively; the percent of Base T in SNPs Mutant is24.2%, and the percent of T into A, C and G is15.70%,69.42%,14.88%respectively; the percent of Base C in SNPs Mutant is25.5%, and the percent of C into A, T and G is17.26%,72.94%,9.80%respectively; the percent of Base G in SNPs Mutant is25.9%, and the percent of G into A, T and C is75.29%,16.60%,8.11%respectively. These results showed that the space mutation factors had thesame effects on the mutation of the A, T, C and G base. The most found base change types are the interchange between A and G, and the interchange between T and C. After comparing the gene sequences oofmutant and wild-type, we found that the mutagenesis of SNPs, InDel and SV have made more than2000genes’ encoding amino acid changed in mutant.3. Genetic analysis and mapping of the mutated gene conferring orange-re d leaves and hullWe found that phenotype of Orange-red trait was controlled by a single recessive allele by comparing the F2plants of the mutant with Chuan Hui907and9311cross. F2population of the cross between mutant and9311was analyzed, and the gene was located on chromosome2, with the genetic distance of0.55cM and1.38cM from SSR marker RM5350and RM12601. Further study was conducted by InDel markers, and the target gene was narrowed down the locus to0.41cM and0.25cM from S1and S2, respectively. After analyzing the300KB sequence attained by the maping analyasis with above molecular markers, we found that there is no available SSR and InDel markers. Therefore, we designed8CAPS markers in this region based on the sequence of indica rice9311, and located the gene into the region of9412bp which only concluding a gene Osl₀6158. Analysis showed that the gene Osl₀6158encoding a R2R3-MYB protein is a regulatory factor,. The reported studies in Arabidopsis and Petunia have showed that R2R3-MYB protein, bHLH protein of MYC family and WD40repeat protein interact with each other to regulate the biosynthetic pathway of anthocyanin in plant.4. Application of mutants in rice breedingThe mutant has good agronomic and economic traits, and specific Orange-red trait that facilitates the selection of rice hybrid plants, so it is a good donor in rice breeding. When hybridization with sterile lines or CMS LINES of different genetic background, we found that the offspring from mutant and Yixiang1A is highly sterile. After hybriding, backcrossing with Yixiang1A, and breeding for many years, we obtained a new hybrid rice CMS line HuaxiangA. Afer hybriding HuaxiangA with different rice restorer lines such as ChuangHui907, ChuangHui4016and ChuangHui1618, we have got new hybrid rice combinations of HuaXiang7, HuaXiang907, HuaXiang4016and HuaXiang1618, which have passed the provincial approval.