利用PCR技术同时检测番茄抗根结线虫基因(Mi-1)和抗叶霉病基因(Cf-9)

Simultaneous identification of multi-genes with resistance to root-knot nematode and tomato leaf mold by PCR reaction in tomato

利用同一PCR反应体系,对分别与番茄抗根结线虫的Mi-1基因和抗番茄叶霉病的Cf-9基因紧密连锁的CAPS(cleaved amplified polymorphic sequences)标记进行同时扩增筛选,扩增的特异性片段与单引物扩增片段吻合.与Cf-9基因紧密连锁的CAPS1标记在抗病试材中可扩增出2775bp的特异片段,且存在TaqⅠ酶切位点,酶切后产生1177bp、446bp、370bp和160bp的不同特异片段;与Mi-1基因紧密连锁的CAPS2为共显性标记,抗性材料中产生915bp的特异片段,TaqⅠ酶切后产生719bp和196bp的特异片段.该体系可用于在同一PCR反应体系中对Mi-1和Cf-92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.

Mi-1 and C f-9 genes in tomato tightly linked with two CAPS （cleaved amplified polymorphic sequence） markers and respectively resistant to root-knot nematode and leaf mold were amplified and screened by using a single PCR reaction. The PCR products were correspond to the amplified bands produced by single CAPS primer. Among them, 2775 bp fragment linked with C f-9 gene was amplified in the resistant tomato lines. The amplified bands were distinguishable after cleavage with the restriction enzyme Taq Ⅰ . Genotype with C f-9 gene could produce 1177 bp, 446 bp, 370 bp and 160 bp bands, respectively. The co dominant CAPS2 marker tightly linked with Mi-1 gene produced 915 bp fragment in the resistant tomato lines. Genotype with Mi-1 gene could produce 719 bp and 196 bp bands, respectively. The replicated stable results proved that two resistant genes could be identified simultaneously by using corresponding CAPS primer under adaptable condition. Compared with single primer PCR, this system is time-saving, labor saving and low cost. It could be very useful for markerassisted selection during early stage in tomato and efficiently speed up breeding procedure.