Development And Validation Of Molecular Markers Linked To The Epistatic Suppressor Gene (esp) Of A Recessive Genic Male Sterility In Brassica Napus L.

The recessive genic male sterility (RGMS) systems have been the important ways for utilizing heterosis in rapeseed because of their stable and complete sterility, extensive distribution of restorers and no negative cytoplasmic effect. At present, there are two RGMS systems mainly used in Brassica napus L. in the world, i.e., digenic RGMS and recessive epistatic genic male sterility (REGMS) which is more successful.9012AB is the type of REGMS. Genetic analysis indicated that the sterility of 9012AB is controlled by two pairs of recessive duplicate genes(ms3ms3ms4ms4) and one pair of recessive epistatic inhibitor gene (espesp). Homozygous recessive sterile gene at both loci can result in male sterility, but the fertility can be restored when the esp gene is in recessive homozygosity or either of the sterile genes is not recessively homozygous. Base on this genetic model, a three-line system for hybrid seed production can be developed in this REGMS system. This system can produce 100% male-sterile population by a temporary maintainer line, and overcome the difficulty of remove 50% male fertile plants during hybrid seed production. It is a trivial and inefficient work to select elite sterile lines and their temporary maintainer lines by using traditional crosses and testcross. Therefore, the most efficient way is to develop molecular markers which tightly linked to the genes (ms3, ms4 and esp) been bred by MAS (Marker Assisted Selection). The purpose of this study is to develop molecular markers tightly linked to the esp gene by using a population which is similar to a segregation F2 population derived from recessive epistatic genic male sterility line 9012AB and its temporary maintainer (T45). Moreover, in order to evaluate the genotype of 22 different rapeseed materials. Genetic analysis on Esp/esp locus was conducted. Based on this result, the genotype of 32 different rapeseed materials were validated by using 44 molecular markers tightly linked to Esp/esp locus developed previously in our lab. The main results were as follows:1. With amplified fragment length polymorphism (AFLP) technology and bulked segregant analysis (BSA),1,024 primer combinations were screened and 5 AFLP fragments (L1, L2, L3, L4, L5) tightly linked to the esp locus were identified. All the markers were located on one side of the esp gene. The genetic distance between the esp gene and the four markers L1, L3, L4 and L5 was the same 0.8 cM, and the genetic distance between the esp and L2 was 2.9 cM.2. L1, one of the four nearest AFLP markers, was sequenced and converted to SCAR marker SCL1 successfully. This SCAR marker was used to analyse in mapping population including 130 individuals. The results showed that the genetic distance between SCL1 and the esp gene was 0.8 cM.3.22 different rape materials with unknown genotype were genetic analyzed in this study, and the genotypes of Esp/esp locus of 17 different rapeseed materials were presumed. Only one has the genotype of EspEsp, and the others have the genotype of espesp.4.44 molecular markers tightly linked to Esp/esp locus developed previously were validated among 32 different rapeseed materials. The results showed that 38 markers were polymorphic, in which the marker genotypes of 12 markers were in completely accord with the genetic genotypes of 27 different rape materials, the conformity accuracy of 5 markers were 96%, and the conformity accuracy of the other markers were range from 11% to 78%.