多花黑麦草抗病基因类似物的克隆及CAPS标记

多花黑麦草具有适应性强、生物产量高和品质好等优点,为我国南方及其他温带地区重要的牧草。本研究利用目前已克隆出的抗病基因(R基因)所编码的蛋白质结构上所具有的共同氨基酸基序NBS-LRR设计寡核苷酸简并引物,对多花黑麦草基因组进行抗病基因类似物(RGA)克隆。从克隆中筛选出具代表性的RGA克隆115个,设计RGA-STS引物113对。利用细胞质雄性不育(CMS)F1群体对RGA-STS引物进行筛选,并采用限制性内切酶酶切扩增多态序列(CAPS)法进行抗病类似物的分子标记,共检测到22对RGA-STS引物的扩增产物具多态性,22个RGA-CAPS标记均被标记于多花黑麦草遗传连锁图中。结果还表明RGA在多花黑麦草遗传连锁图中分布范围较广,并具有簇状分布特性。

Isolation and cleaved amplified polymorphic sequence markers of disease resistance gene analogues in Lolium multiflorum

Italian ryegrass(Lolium multiflorum) is one of the most important temperate forage grasses.It is the most widely cultivated annual forage grass in China due to its high forage yield and good quality.The commonest class of plant disease resistance genes cloned so far belongs to the NBS-LRR group which contains nucleotide-binding sites(NBS) and a leucine-rich repeat(LRR).In this study,degenerate oligonucleotide primers were designed from the conserved motives of cloned disease resistance genes and used for a polymerase chain reaction(PCR) to isolate disease resistance gene analogs(RGAs) from the genome of Italian ryegrass.One hundred and fifteen representative RGAs were selected,and 113 pairs of specific sequence tagged site(STS) primer sets were designed.The F_1 population of cytoplasmic male sterility(CMS) containing 124 individuals,and 18 endonucleases were used to map twenty-two cleaved amplified polymorphic sequence(CAPS) markers to the genetic linkage map of Italian ryegrass constructed mainly by SSR markers.The results indicated that RGAs are distributed in all seven genetic linkage groups of Italian ryegrass and that some RGAs are clustered.