Finning Mapping Of The Brown Planthopper Resistance Gene Bph14 And Identification And Analysis Of The Sucrose-starvation Sensitive-1 Sss1 Mutant In Rice

In this study, by using the classic map-based cloning strategy we successfully delimited the Bph14 locus to a 34-kb region based on the result of the rough mapping previously performed in our lab, the two powdery mildew resistance protein PM3b gene that predicted by TIGR were taken as the candidate gene for Bph14 based on the analysis for the prediction of the candidate gene in the putative target interval. In the study of the identification and analysis of the sucrose-starvation sensitive-1 sssl mutant in rice, based on the result of the gene mapping and the effect of the exogenous sugar on the mutant, it was preliminary supposed that the phenotype of the mutant is owed to the lack of sucrose which results from the mutant in the sucrose synthesis gene flanking the marker that closely linked to the sssl.RI35 and RI113, selected to make new populations for fine scale and high resolution of target QTL of Bph14, in which the two minor QTLs confirmed by the analysis of the genetic background. In the case of the selection of the most desirable plants with the only target QTL of Bph14 in the TN1/RI35 and TN1/RI113 RIL population,533 plants achieved from about 5000 progenies of the 298 recombinants of the TN1/RI35 and TN1/RI113.A linkage analysis, mainly employing microsatellite markers, was carried out through bulked segregant analysis and recessive class analysis (RCA), and the marker RM1221 that closely linked to Bph14 was screened. After achieving the marker RM1221 that closely linked to Bph14, the genotyping evaluation of the 533 plants with the only target QTL of Bph14 was performed using the markers that flanking the marker RM1221 and the genetic map of the target interval was constructed. Combination the evaluation of the BPH resistance and the genotyping, the Bph14 locus was narrowed-down to the interval between the markers RM570 and SM4. RT12-5, having a double crossover event in the interval between the markers RM514 and SM4, was selected from the 533 plants with the only target QTL ofBphl4.Combination the evaluation of the BPH resistance and the genotyping of the self-pollinated progenies of RT12-5, the Bphl4 locus was confirmed in the interval between the markers RM1221 and SM4. Among about 3000 self-pollinated progenies, one plant having a recombination event in the interval between the markers RM1221 and 76-2, another plant having a recombination event in the interval between the markers G1318 and SM4 was screened. Combination the evaluation of the BPH resistance and the genotyping of the two recombinants, the Bph14 locus was further narrowed-down to the interval between the markers 76-2 andG1318.Based on the information of the physical map that constructed previously in our lab, the physical distance between 76-2 and G1318 was about 34 kb, thus the Bphl4 locus was delimit to a 34-kb region. Combination the candidate gene prediction performed by GAAS,TIGR and NCBI and the information and the characterization of the cloned resistance gene, the two powdery mildew resistance protein PM3b gene that predicted by TIGR were taken as the candidate gene for Bph14.One useful natural mutant isolated and characterized. Genetic analysis showed that the mutant was controlled by a single recessive gene. In M3 generation, there was no segregation of any trait within the M3 families when they descended from M1 individuals. One sucrose synthesis gene flanking the marker that closely linked to the sssl was taken as the putative gene of the sssl. In the mutant grown on nutritionally deficient media experiment, the root growth of the mutant rescued when the mutant grown on the 1/2 MS media containing sucrose, however, the root growth of the mutant didn’t rescue when the mutant grown on the 1/2 MS media minus sucrose, showing that the mutant possibly defective in sucrose synthesis or metabolism and sugar-response. Owing to the sensitivity to sucrose starvation, the mutant thus designated as sssl (sucrose-starvation sensitive 1).In the mutant grown on nutritionally deficient media experiment, the nutritionally deficient media prepared by minus only a single ingredient of the 1/2 MS media. The root growth of the mutant rescued in concentration-dependent manner when germinated seeds were plated in the distilled water supplemented with various concentrations of sugar. The result showed that the other ingredient in the 1/2 MS media didn’t have effect on the root growth of the mutant and more directly revealed that the phenotype of the mutant is owed to the lack of sugar. As the product of photosynthesis and the sucrose hydrolysis respectively, Suc(sucrose), Glu(glucose) and Fru (fructose), can rescue the root growth of the mutant, but Mtl(mannitol) or starch, the precursor of Sue, can’t rescue the root growth of the mutant.Suc, Glu and Fru levels have the similar effect on the seedling growth of the mutant when the mutant grown on the 1/2 MS media containing sugar or germinated seeds plated in the distilled water supplemented with sugar. When the mutant grown on the 1/2 MS media containing Suc, but not Glu or Fru, at the same time when germinated seeds plated in the distilled water supplemented with Suc, but not Glu or Fru, the tiller buds initiated and the tillers outgrown. The fact that Suc and Glu and Fru have different effect on the tiller buds initiation and the tillers outgrowth preliminary revealed that the phenotype of the mutant is owed to the lack of sucrose. Meanwhile, for the first time the study on the mutant revealed that the sucrose can act by affecting the tiller buds initiation and the tillers outgrowth.