【作者】 李君明； 宋燕； 陈丽静； 杨宇红； 徐和金； 冯兰香； 张智； 周永健；

【Author】 Li Junming Song Yan Chen Lijing Yang Yuhong Xu Hejin Fen lanxiang Zhangzhi and zhou Yongjian (Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081 )

【机构】 中国农科院蔬菜花卉研究所；

【摘要】 本文利用同一PCR反应体系,对分别与番茄抗根结线虫的Mi基因和抗番茄花叶病毒病的Tm22基因紧密连锁的SCAR标记进行了同时扩增筛选,扩增的特异性片段与单引物扩增片段基本完全吻合,其中与Mi基因紧密连锁的SCAR1标记为共显性标记,抗感试材均产生750bp的特异片段,纯合和杂合抗病基因型存在TaqI酶切位点,酶切后分别产生了570bp、200bp和750bp、570bp、200bp的不同特异性片段,而感病基因型试材无酶切位点;与Tm22因紧密连锁的SCAR2标记也为共显性标记,抗感试材均产生950bp的特异片段,杂合抗病基因型和感病基因型有HindIII酶切位点,杂合抗病材料为950bp和490bp片断;感病材料为490bp;纯合抗病基因型无HindIII酶切位点,酶切结果仍为950bp产物。经反复验证,结果十分稳定、准确可靠,可用于在同一PCR反应体系中对2个抗病基因进行同时筛选鉴定。该体系的建立不仅省时、省工、节省费用,而且可用于苗期早期辅助选育,加快番茄育种进程。更多还原

【Abstract】 Single PCR reaction with two SCAR markers respectively tightly linked with M and Tm22genes in tomato, which resistant to root-knot nematode and Tomato mosaic virus, has been used to screen the multiplex bands. The PCR products were almost completely correspond to the amplified bands produced by single SCAR primer. Among them, codominant SCAR1 marker tightly linked with M gene produce 750bp fragment in both resistant and susceptible tomato lines. The amplified bands from susceptible and resistant lines were distinguishable after cleavage with the restriction enzyme TaqI. Genotype with homozygous and heterozygous Mi gene could produce respectively 570bp, 200bp and 750, 570bp, 200bp bands. Susceptible genotypes still present 750bp fragment. The dominant SCAR2 marker tightly linked with Tm22gene would produce 950bp fragment in both resistant and susceptible tomato lines. The amplified bands from susceptible and heterozygous were distinguishable after cleavage with the restriction enzyme HindIII. Genotype susceptible and heterozygous with Tm22 gene could produce respectively 490bp and 950,490bp bands, homozygous genotypes still present 950bp fragment. The replicated stable results proved that two resistant genes could be identified simultaneously by using corresponding SCAR primer under adaptable condition. This system compared with single primer PCR would be time saving, less labor and low cost. It could be very useful for marker-assisted selection during early stage in tomato and efficiently speed up breeding procedure.更多还原