旋毛虫成虫cDNA文库的建立及PCNA基因克隆和序列分析

Molecular cloning and sequencing of Trichinella spiralis PCNA antigen

目的建立旋毛虫成虫cDNA文库,对旋毛虫增殖细胞核抗原PCNA基因进行克隆和测序分析。方法收集纯净的旋毛虫成虫,提取RNA,构建cDNA文库。根据GenBank中登录的PCNA基因的序列设计并合成引物,用PCR方法从旋毛虫成虫基因库筛选PCNA基因;将PCR产物与pUM-T载体连接,转化到感受态细胞DH5α,进行测序和同源性比较。结果成功构建旋毛虫成虫cDNA文库,克隆PCNA基因序列与GenBank登录的PCNA基因同源性为94%。结论成功构建了旋毛虫成虫cDNA文库;成功克隆了PCNA基因。

Objectives To construct a cDNA library of Trichinella spiralis and clone and sequence T. spiralis PCNA.Methods Adult T. spiralis worms were collected, their RNA was extracted, and a cDNA library was constructed.Primers were designed and synthesized in accordance with the PCNA gene sequence in GenBank. Polymerase chain reaction （PCR） was used to screen genes of adult T. spiralis worms for the PCNA gene. PCR products were ligated to pUMT and transformed into DH5α competent cells, and sequences and homology were compared. Results A cDNA library of the adult T. spiralis worm was successfully constructed and the PCNA sequence was cloned; its identity with the sequence in GenBank was 94~. Conclusion A cDNA library of the adult T. spiralis worm was successfully and the PCNA gene was successfully cloned.