PLUG引物和IT引物在顶芒山羊草分子标记建立中的通用性分析

Universality Analysis of PLUG and IT Primers in Establishing Molecular Markers of Aegilops comosa

分子标记是检测和追踪小麦背景中近缘物种染色体的重要手段。依据禾本科物种基因共线性原则,基于基因序列设计的引物在不同物种中可能具有通用性。本研究对水稻PLUG引物和簇毛麦IT引物在顶芒山羊草中的通用性进行了分析,并建立了顶芒山羊草染色体特异IT标记。供试的397对PLUG引物和841对IT引物对小麦-顶芒山羊草双二倍体与中国春的扩增结果显示,377对(95.0%)PLUG引物和593对(70.5%)IT引物可以在供试材料中扩增出清晰条带。因此,PLUG引物在顶芒山羊草中的通用性优于IT引物。相比小麦对照,PLUG引物和IT引物能在小麦-顶芒山羊草双二倍体中扩增出多态性条带的引物数分别有17对和5对,分别占供试PLUG引物和IT引物的4.3%和0.6%。利用获得的IT多态性引物对一套小麦-顶芒山羊草附加系和一套小麦-卵穗山羊草附加系进行扩增,结果发现仅引物CINAU1060、CINAU1465和CINAU1517能在相应附加系中扩增出大小分别约为300、480bp和320bp的多态性标记,为小麦背景中顶芒山羊草染色质的鉴定提供了新的检测手段。然而,本研究中IT引物建立顶芒山羊草分子标记的比率仅为0.4%,低于我们之前用PLUG引物建立顶芒山羊草分子标记的比率(8.9%)。

Molecular marker plays an important role in detecting and tracking chromosomes of related species in wheat background. According to the principle of gene collinearity in Gramineae species, primers designed based on gene sequences might be universal in different species. In this research, the universality of PLUG primers for Oryza sativa and IT primers for Dasypyrum villosum were analyzed in Aegilops comosa , and chromosome specific IT markers of Ae.comosa were established. The amplification results of 397 pairs of PLUG primers and 841 pairs of IT primers in wheat- Ae.comosa amphiploid and Chinese Spring showed that 377 pairs (95.0%) of PLUG primers and 593 pairs (70.5%) of IT primers could amplify clear bands in the tested materials. Therefore, the universality of PLUG primers in Ae.comosa was better than that of IT primers. Compared with Chinese Spring, the number of PLUG and IT primers that could amplify polymorphic bands in wheat- Ae.comosa amphiploid was 17 pairs and 5 pairs, which accounted for 4.3% and 0.6% of the PLUG and IT tested primers, respectively. In this research, a set of wheat- Ae.comosa addition and a set of wheat- Ae. geniculata addition were amplified with the obtained IT polymorphic primers. The results showed that 300, 480 bp and 320 bp in length polymorphism bands could be amplified by primers CINAU1060, CINAU1465 and CINAU1517 in relative additions, respectively, which provided new method for the identification of Ae.comosa chromatin in wheat background. However, the ratio for molecular marker development of Ae.comosa using IT primers was 0.4%, which was much lower than that of PLUG primers (8.9%).