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利用雄性不育系培育新品种和生产杂种一代,在产量、品质等方面均具有独特优势。前期课题组已通过多代连续杂交获得丹参雄性不育与可育近等基因系。该文在前期工作基础上采用AFLP和BSA技术对378对引物进行筛选,发现7对引物和26个标记与丹参育性调控基因紧密连锁,由此构建出连锁遗传图谱。以前期发现的E11/M4-208标记为模板,运用染色体步移技术,在不育和可育系中分别扩增出2 028,2 027 bp的差异片段,发现4个突变碱基位点均处于内含子中。在所有差异标记中发现E01/M09-418,E05/M13-308,E05/M04-750,E01/M01-204 4个与丹参育性调控基因紧密连锁的标记,与拟南芥1,3,5号染色体相似性达100%。其中与育性调控基因距离很近(2.1 c M)的E01/M09-418与同处于拟南芥第3号染色体的核不育基因MS2相似度高达100%。不育系中获得的2 028 bp片段与MS2基因在两处相似度达100%。与拟南芥第5号染色体相似度100%的E05/M04-750,与杨树POP085-M05基因和拟南芥中低亲和力钙逆向转运蛋白序列具有较高相似性,且E非常低。该文遗传图谱的构建和功能性差异序列的发现,为后续准确锚定丹参基因组中育性调控基因及揭示其功能奠定了坚实基础。
The use of male sterile lines to cultivate new varieties and hybrid generation, in terms of yield, quality and so has a unique advantage. Previous research group has been obtained by multiple generations of Salvia miltiorrhiza male sterile and fertile near isogenic lines. Based on the previous work, 378 pairs of primers were screened using AFLP and BSA techniques. Seven pairs of primers and 26 markers were found to be closely linked to the fertility regulation gene of Danshen, and the linkage map was constructed. Previously found E11 / M4-208 markers as a template, the use of chromosome walking technology, respectively, in sterile and fertile lines were amplified 2,028,2 027 bp fragments were found in all four mutant base sites In intron. Four markers (E01 / M09-418, E05 / M13-308, E05 / M04-750, E01 / M01-204) closely linked to the fertility regulation gene of Salvia miltiorrhiza were found in all the differential markers, The chromosome 5 similarity of 100%. Among them, E01 / M09-418 which is close to the fertility gene (2.1 c M) has a similarity of 100% with MS2 which is the same as the male sterility gene MS3 in Arabidopsis thaliana. The 2 028 bp fragment obtained from the CMS line was 100% identical to the MS2 gene at two sites. E05 / M04-750, which is 100% similarity to Arabidopsis chromosome 5, has high similarity to the poplar POP085-M05 gene and Arabidopsis low-affinity calcium antiporter sequence, and E is very low. The construction of the genetic map and the discovery of functional differential sequences laid a solid foundation for the subsequent accurate anchoring of fertility-controlling genes in Salvia miltiorrhiza genome and revealing its function.