番茄抗番茄花叶病毒和斑点萎凋病毒病基因PCR标记的同时鉴定

Simultaneous Identification of Multi-Genes with Resistance to Tomato Mosaic Virus and Tomato Spotted Wilt Virus by PCR Markers in Tomato

利用同一PCR反应体系,对分别与番茄的抗番茄花叶病毒病的Tm22基因和抗斑点萎凋病毒病的Sw-5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完全吻合,其中与Tm22基因紧密连锁的SCAR1标记为共显性标记,抗感试材均产生800bp的特异片段,杂合抗病基因型和感病基因型有HindIII酶切位点,酶切结果为:纯合抗病RR:950bp;杂合抗病Rr:950bp+500bp+300bp+150bp;感病的rr:500bp+300bp+150bp,纯合抗病基因型无HindIII酶切位点。与Sw-5基因紧密连锁的SCAR2标记为显性标记,只有抗病试材扩增出400bp的特异性片段。经反复验证,结果稳定、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。

Single PCR reaction with two SCAR markers, respectively, tightly linked with Tm22 and Sw-5 genes intomato, which resistant to tomato mosaic virus and tomato spotted wilt virus, has been used to screen the multiplex bands.The PCR products were completely correspond to the amplified bands produced by single SCAR primer. Among them,codominant SCAR1 marker tightly linked withTm22 gene produced 950bp fragment in both resistant and susceptible tomatolines. The amplified bands from susceptible and heterozygous were distinguishable after cleavage with the restrictionenzyme Hind III. Genotype susceptible and heterozygous with Tm22 gene could produce respectively 500,300,150 and 950,500,300 ,150bp bands. homozygous genotypes still present 950bp fragment. The dominant SCAR2 marker tightly linkedwith Sw-5 gene would produce only 400bp PCR product in resistant genotype. The replicated stable results proved thattwo resistant genes could be identified simultaneously by using corresponding SCAR 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 marker-assisted selection during early stage in tomato and efficiently speed up breeding procedure.