分子标记鉴定EMS诱变高粱突变体的真伪

Identification the Authenticity of EMS Mutagenic Mutants by Molecular Markers of Sorghum

甲基磺酸乙酯(Ethylmethylsulfone,EMS)作为简易、有效的诱变剂在高粱诱变育种中得到广泛应用。由于诱变后代开放授粉,易接受其它高粱材料花粉出现假突变体的可能,因此开发一套实用的分子标记鉴定突变体真伪是非常有必要的。研究基于26份高粱材料的重测序数据,共开发13390个≥3 bp重复的SSR分子标记。在基因上和非基因上各选择100个标记进行有效性和丰度检测,最终选择PCR扩增目标条带较单一、多态性丰富的30个标记用于EMS突变体真伪鉴定。选择5个与BTx623杂交获得的RIL群体株系、5个已知BTx623 EMS诱变突变体以及5个新筛选的BTx623 EMS诱变突变体为对象,检测30个SSR分子标记的有效性及鉴别能力。与亲本BTx623相比,5个RIL株系中PCR扩增条带存在差异的标记分别为10、11、12、10和9个。而5个已知突变体以及新筛选突变体3T133、4A317和2E52 PCR扩增条带与亲本BTx623一致。新筛选突变体4A478和4A211分别有6个和5个标记PCR扩增条带大小与亲本BTx623存在差异,结合田间表型,判定两者皆为假突变体。由此可见,本研究筛选鉴定的≥3 bp重复的30个SSR分子标记可以用于EMS突变体真伪鉴定。

EMS(Ethylmethylsulfone),as a simple and effective mutagen,has been widely used in sorghum mutation breeding.Because of the mutation offspring open pollination,it is possible to produce pseudo-mu-tants by crossing with other sorghum materials.Therefore,it is absolutely necessary to develop a set of useful molecular markers for identifying the authenticity of mutants.Based on the resequencing data of 26 sorghum materials,13390 SSR markers with≥3 bp repeats were developed.100 SSR markers were selected on gene and non-gene respectively to test the effectiveness and abundance of markers.Finally,30 SSR markers with relatively single PCR target bands and rich polymorphisms were selected for identification the EMS mutants.Five RIL lines which were BTx623 crossing with other different sorghum materials,five known mutants from BTx623 treating with EMS and five newly screened mutants were selected to test the effectiveness and discrimi-nation ability of 30 markers.Compared with BTx623 and five RIL lines,10,11,12,10 and 9 markers had different PCR amplification bands,respectively.The PCR amplification bands of known mutants and newly screened mutants 3T133,4A317 and 2E52 were consistent with BTx623.Compared with BTx623,6 and 5 SSR markers had different PCR amplification bands of newly screened mutants 4A478 and 4A211,respective-ly,which were judged as pseudo-mutants combined with field phenotype.Thus,30 SSR markers with≥3 bp repeats can be used for identifying the authenticity of EMS mutants in this study.