与多花黑麦草抗灰叶斑病相关基因紧密连锁的RGA-CAPS标记筛选

以多花黑麦草抗灰叶斑病品种Sach iaoba、感病品种M inam iaoba及其杂种F1分离群体为材料,采用接种鉴定和分群分析法(BSA)进行抗病基因类似物限制性内切酶酶切多态性(RGA-CAPS)筛选。得到与多花黑麦草抗灰叶斑病相关基因紧密连锁的RGA-CAPS标记,该标记与灰叶斑病抗性相关基因紧密连锁,连锁距离为4.94cM。测序和BLASTX查询结果显示,该RGA与水稻的1个NBS-LRR类抗性蛋白(Genbank/NCB I:AAT81729.1)同源性高达67%,序列分析结果显示,对应于抗、感病性的该RGA片段长度均为361 bp,两片段之间存在多个单核苷酸差异(SNPs)。利用SNPs重新设计引物,并应用新设计引物成功地将RGA-CAPS标记转换成共显性STS标记。

A RGA-CAPS Marker Closely Linked to Grap Leaf Spot Resistance-related Gene of Lolium multiflorum

A two-way pseudo-testcross F_1 population consisting of 162 individuals was composed by crossing gray leaf spot resistant individual (Sachiaoba) and sensitive one (Minamiaoba). One resistance gene analogs-cleaved amplified polymorphic sequence (RGA-CAPS) marker linked to gray leaf spot resistance gene of Italian ryegrass was developed by the method of bulked segregant analysis (BSA). Linkage analysis indicated that the RGA-CAPS marker was tightly linked to the gray leaf spot resistance related gene, and genetic distance was 4.94 cM. Sequencing of the RGA fragments indicated that the two fragments related to gray leaf spot disease resistant and sensitive were the same size, 362 bp, but there are several SNPs between them. According to the SNPs, the RGA-CAPS marker was converted to a co-dominant STS marker. BLAST analysis revealed that the RGA was closely homologous to a NBS-LRR type protein of rice, accession code: AAT81729.1.