Screening Of Molecular Markers Linked To Boron Efficient Genes And Gene Mapping In Brassica Napus

Boron (B) is an essential nutritional element for plant growth and development. Rapeseed (B. napus) usually requires a great deal of boron and is very sensitive to B deficiency. Rapeseed will result in "flowering without seed" when it suffers from B deficiency. The main producing areas of B. napus in China are deficient or extremely deficient in soil available boron. The main means to resolve boron deficiency is to increase the rate of boron fertilizer, whereas boron is a non-renewable mineral resource. Considerable genotypic variations in response to B deficiency exist among rapeseed varieties, so our research group has been screening high boron efficiency germplasm rapeseed resources since 1991 and also conducted study on physiological mechanism of rapeseed in response to boron deficiency, and mapping the boron efficiency genes. The B efficiency was proved to be controlled by QTL in B. napus, which consist of one major QTL and three micro QTLs. To fine mapping and clone these genes, this study was involved in screening the linked markers of the boron efficiency genes, by using BC₁ population constructed by a new variety being more sensitive to boron deficiency and the former boron-efficient variety in B. napus. And we also design some gene-specific primers for mapping of the B-efficient functional markers, by using sequence of the B-efficient gene related to B uptake and translocation in Arabidopsis and Nicotiana tabacum.1. Constructed BC₁ population by using a new B-inefficient variety Westar 10 as the recurrent parent and a B-efficient variety Qingyou 10 as the donor parent. The growth of BC₁ population was investigated under low B level (B=0.25μM) with solution culture. The results showed significant segregation in the phenotype growth among the individuals.2. Constructed B-efficient DNA Bulk and B-efficient DNA Bulk by 10 B-efficient individuals and 10 B-inefficient individuals, respectively. For screening molecular markers linked to the B-efficient genes, BSA was applied combining with AFLP. By analysis of 1124 pairs of AFLP primer, we finally got 23 linked markers. These markers were used to constructed a genetic linkage map for targeted region, two overlapped QTLs which associated with plant biomass and B-deficient symptom were detected in this region.3. Mapping the linked markers in BQDH and TNDH population, theses markers were eventually located in 4 linkage groups in BQDH population involving A4, A6, C8, C9, and A2, A4, C3, C8 in TNDH population.4. Comparative mapping was carried out between the targeted region in B.napus and genome of Arabidopsis. The targeted region of A6 and C9 in BQDH population corresponded to the A and D region of chromosome 1 in Arabidopsis, the region of A4 in both BQDH and TNDH population corresponded to chromosome 5 in Arabidopsis; Analyze the sequence of linked markers by BLAST, the corresponding homologous sequence distributed in all of 5 chromosomes in Arabidopsis, so these markers may be linked to the genes concerning B transporters and B channel to some extent.5. Homologous sequences of markers linked in Brassica species were found and the flanking sequences were used for primer design. By this way, two linked AFLP markers were transformed to simple specific SSCP markers successfully.6. Nine functional markers were developed from homologous sequences of genes related to boron efficient uptake and transport in Arabidopsis thaliana and Nicotiana tabacum, and eight of them were finally mapped in A1, A3, A7, C3 and C6 of BQDH genetic map respectively.