Marker development and gene identification for blackleg (Leptosphaeria maculans) disease resistance in canola (Brassica napus).

摘要

A consensus ultradense genetic recombinant map was constructed using sequence related amplified polymorphism (SRAP) markers and used to tag a blackleg disease resistance gene LepR3 in 'Surpass 400'. Marker development was also performed using comparative genomic sequencing to find single nucleotide polymorphism (SNP) markers for the other two disease resistance genes Rlm1 and Rlm3 in 'Quinta' and 'Glacier', respectively. The objective of gene mapping is to use molecular markers linked closely to disease resistance genes for marker assisted selection (MAS) in cultivar development and for pyramiding different resistance genes into a cultivar to improve the degree of resistance. Furthermore, cloning of these disease resistance genes will be essential to fully understand the underlying mechanism of disease resistance and to manipulate disease resistance genes for managing the disease effectively.;The dominant resistance gene LepR3 in 'Surpass 400', introduced from B. rapa subsp. sylvestris through gene introgression, was targeted for marker development and gene cloning. To use the ultradense genetic recombination map, the same primer combinations for the map construction were used to find the common SRAP markers that were linked to the disease resistance gene LepR3 in the segregating DH line population of 'Westar' and 'Surpass 400'. The integration of these SRAP markers allowed the use of SRAP markers on the map to find SRAP markers linked more closely to LepR3. With 384 primer combinations, two SRAP markers, R269 and G278 linked to LepR3 were developed and R269 was found to correspond to SRAP marker 1217Ar269 on the N10 linkage group, which contained 508 markers.;The SRAP markers on the N10 linkage group were selected to screen the gene tagging population and three markers, 210Ay442, 0127Fr382 and 1128BG275, were found to co-segregate with the gene LepR3. After analysis of these SRAP markers with the population of 3,900 plants, 52 recombinant plants between SRAP markers 1217Ar269 and 0127Fr382 were selected for further analysis and the region containing the gene LepR3 was identified. Moreover, SRAP marker 0127Fr382 was found to be the closest marker to the resistance gene LepR3 with a genetic distance of 0.3 cM. (Abstract shortened by UMI.);An ultradense genetic recombinant map was developed with a 58 DH line population of 'Westar x Zhongyou 821' and in total 13,551 SRAP markers were integrated on 19 linkage groups that corresponded to the 19 chromosomes in B. napus. All SRAP markers were generated with a total of 1,634 primer combinations including 12 fluorescently labeled primers and 442 unlabeled ones. These 13,551 markers were put into 1,055 bins, resulting in a map length of 1,604.8 cM. Furthermore, all 19 linkage groups were assigned to the previously reported N1--N19 linkage groups of B. napus by integrating 55 microsatellite or simple sequence repeat (SSR) markers that had been used to construct previous maps in this species.