Identification Of The S Haplotypes In Cabbage (Brassica Oleracea L.var.capitata)

Cabbage （Brassica oleracea L. var. capitata） is a typical cross-pollinated crop with obviousheterosis. Self-incompatible line is one of the two main ways in utilizing heterosis in B. oleracea.Because the crossed two materials carry the same S haplotype, seed setting happens hardly resultingfrom the incompatibility response. Therefore, it is very important for cabbage breeding to identifyrapidly and accurately the S haplotypes of breeding materials. The conventional method of test-corssingis highly time-consuming and the results are distorted sometimes by environmental factors. Withpublished S haplotypes increasing, rapid identification of cabbage S haplotypes can be achieved viaDNA sequencing and blast analysis. The results will not be influenced by environmental factors, andhave high accuracy. Identification of S haplotypes will provide guiding for making the combinationsand avoid the incompatibility response between parents.SRK and SCR/SP11are the determinants of stigma and pollen in the SI recognition response,respectively. By PCR amplification of SRK and ClassⅡSCR/SP11with specific primers, sequencingand blast analysis, molecular identification of S haplotypes of140accessions, most of which arecabbage inbreed lines developed from cultivars, has been carried out in this study. The results ofmolecular identification were confirmed by fluorescence microscopy identification in part of thematerials. The main results are as follows:1. Types and distribution of S haplotypes in cabbagesIn the140cabbage accessions,8materials of ClassⅠwere not indentified by sequencing analysis,and the other132materials were identified as16S haplotypes. The distribution of S haplotypes are17S₂haplotype,17S₅haplotype,3S₆haplotype,13S₇haplotype,1S₁₃haplotype,5S₁₄haplotype,25S₁₅haplotype,1S₁₆haplotype,3S₂₃haplotype,5S₂₈haplotype,5S₃₃haplotype,1S₃₅haplotype,8S₄₅haplotype,4S₅₀haplotype,3S₅₇haplotype and3S₆₈haplotype. The frequency of different Shaplotypes in this study range from0.71%to20.71%, and S₁₅was the most common haplotype, whileS₁₃, S₁₆and S₃₅were the lowest frequency haplotypes.In this study, S₆₈is the new identified S haplotype of cabbage. So far, the number of S haplotypesin cabbage increased to26. SRK kinase domain （Exon2to5） sequences of S₁₆、S₃₅、S₄₅、S₅₀and S₅₇wereobtained.Three pairs of primers for SCR/SP11were developed for rapidly screening the S haplotypes ofClassⅡ in cabbage. S₂and S₅haplotyopes could be amplified with BoSC2F1/BoSC2R1, while S₂haplotype could be distinguished with BoSC2F2/BoSC2R2, and S₁₅haplotyope could be identified onlywith S₁₅SPF/S₁₅SPR. The three pairs of primers are applicable to the identification of S haplotypes inthe ClassⅡinbred lines of cabbage.2. Identification of fluorescence microscopyIn order to further verify further the results of molecular identification,51cross-combinationsamong14S haplotypes were identified by fluorescence microscopy. The results showed that incompatibility or weak incompatibility happened between two materials with same S haplotype, whilecompatibility was detected between two materials with different S haplotypes. The results wereconsistent with molecular identification of S haplotypes.