Generation And Characterization Of Bentazon Susceptible Mutants Of Rice And Molecular Characterization Of CYP81A6Gene

High purity of hybrid seeds is one of crucial factors for the successful adoption of hybrid rice technology. The ever-changing climate and environmental conditions, and the genetic make-up of male sterile lines are the major factors that are responsible for the incompleteness of male sterility in hybrid seed production. The recessive bentazon susceptibility trait in rice was bred into male sterile lines; thus seedlings of self-pollinated male sterile lines could be easily removed by a spray of bentazon onto seedlings in hybrid rice production. Two such male sterile lines, one CMS line and one P/TGMS line, which are otherwise excellent, were subjected to gamma-ray mutagenesis for bentazon susceptibility. Three bentazon susceptible mutants were selected and their genetic and agronomoic characteristics were investigated. The usefulness of bentazon susceptibility as a marker in hybrid rice production was demonstrated using the new male sterile line. Furthermore, the molecular characteristics of bentazon resistant gene(CYP81A6) were investigated.The results were as follows:1. A cost-effective mutant selection system was developed with～20fold reduction of workload; three betazon susceptible mutant lines of commercial male sterile lines were developed; herbicide test proved that all bentazon susceptible seedlings of three mutants could be completely killed at a concentration as low as400mg/1.2. The three bentazon susceptible mutant lines were identified to carry independent mutations of the CYP81A6gene with deletions of lbp,2bp and11bp. All deletions would result in truncated and dysfunctional CYP81A6enzymes. Two molecular markers were also developed for latter two mutations, which are expected to facilitate the efficient use of the mutations in future breeding programs.3. No significant negative effects of bentazon susceptibility on other agronomic traits of both the mutants and their F1hybrids with other varieties were observed, which indicated that the mutant lines could directly replace their parental lines in commercial hybrid rice production.4、To determine the expression pattern of CYP81A6, transgenic rice plants with a CYP81A6promoter-GUS fusion gene were produced and the GUS activity was examined for different tissues of the transgenic plants. The results showed that the promoter of CYP81A6was constitutive promoter. To determine the sub-cellular localization of CYP81A6, CYP81A6was fused with yellow fluorescence protein gene and the heterologous gene was transiently expressed in rice protoplasts, the results demonstrated that CYP81A6protein was localized in ER.