接头连接介导的PCR 扩增马铃薯抗晚疫病基因侧翼序列研究

利用接头连接介导的PCR 技术，以前期NBS profiling 试验获得的1 条马铃薯抗晚疫病基因
片段为基础，设计巢式基因特异性引物，扩增获得两侧序列。结果表明：基因组DNA 经DraⅠ酶切后，
在已有536 bp 片段的左侧和右侧均得到序列延伸，右侧扩增得到2 301 bp，左侧扩增得到820 bp，去除
边界序列后，向左延伸789 bp，向右延伸2 273 bp，拼接后共长3 443 bp。采用GENSCAN 进行基因预测，
发现包含内含子在内的晚疫病抗性基因全长2 413 bp，在该预测基因 的UTR 区域设计特异性引物，在马
铃薯抗病和感病的基因型中扩增包含完整基因在内的2 571 bp 序列，发现该特异候选基因与晚疫病抗性相
连锁。

Studies on Flanking Sequence of Potato Late Blight Resistance Gene by Adapter Ligation-mediated PCR Amplifying

Based on a potato（Solanum tuberosum L.）late blight resistance gene fragment
obtained by NBS profiling，specific nested primers were designed to amplify flanking sequences of the
fragment using adapter ligation-mediated PCR. The results indicated that，after digestion of genomic
DNA by restriction enzyme Dra Ⅰ，2 301 bp and 820 bp length sequence was amplified at the right and
left side of the known gene 536 bp fragment，respectively．Finally，the fragment was elongated 2 273
bp at right and 789 bp at left，which assemble a 3 443 bp full length sequence. A 2 413 bp candidate
gene including intron and extron was predicted with the software GENSCAN．A primer，RDZ，specific
to UTR region of the gene，was developed to amply resistant and susceptible genotypes．The results
showed a 2 571 bp fragment amplified by RDZ is linked to late b light resistance.