褐飞虱取食水稻幼苗cDNA文库的构建及筛选

Construction and screening of a cDNA library from rice seedlings exposed to brown planthopper

摘要: 以褐飞虱取食 3 2 h的水稻幼苗为材料构建了 c DNA文库 ,初始文库含 3 .2× 1 0 6 个克隆 ,重组率为 86%,取 1 .0× 1 0 6个克隆子扩增一次 ,收集到 80 m L滴度为 1 .2× 1 0 9pfu/ m L的扩增文库 .随机挑取 2 0个克隆以 T7和 M1 3 reverse为引物进行 PCR扩增 ,以鉴定插入片段的大小 ,结果发现多数片段的长度在 1～ 4.0Kb左右 ,少数片段在 4Kb以上 ,个别片段在 6Kb左右 .以在受褐飞虱取食的水稻幼苗中特异表达的 ESTBp Hi0 0 8为探针 ,筛选 c DNA文库 ,得到该基因的 c DNA全长

Abstract: cDNA library was constructed using rice seedlings exposed to BPH for 32 h as materials. The primary library contains 3.2×10~6 clones. The rate of recombination is 86%. 1.0×10~6 clones were amplified, and 80 mL amplified library was obtained. The titer of amplified library is 1.2×10~9 pfu/mL. To estimate the length of inserts, 20 clones were selected randomly and analyzed by PCR method using T7 and M13 reverse primer. The results demonstrated that the size of most inserts is 1～4 Kb, some inserts is larger th...