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目的 克隆日本血吸虫中国大陆株 2 0 .2 k Da分子的编码基因。方法 根据筛选到的日本血吸虫肝期童虫 c DNA文库阳性克隆之一的插入片段序列 ,设计合成 1对引物 ,通过 PCR技术对其最大开放阅读框进行扩增 ,然后克隆入 p GEM- T载体 ,并对克隆产物进行核苷酸序列测定和分析 ,利用在线软件推测其编码氨基酸的序列。结果 利用 PCR方法从上述插入片段中扩增出 1条单一的 DNA条带 ,大小介于 5 15 bp与 6 97bp之间。将此产物克隆入 p GEM- T载体 ,双酶切和 PCR反应结果都出现与前述结果一致的目的片段 ;核苷酸序列测定结果表明此开放阅读框全长 5 6 4bp,在线软件分析结果表明 ,此片段编码 187个氨基酸 ,编码肽段分子量大小约为 2 0 .2 k Da。结论 本次实验成功扩增和克隆出目的基因片段 ,为下一步研究奠定了基础
Objective To clone the gene encoding 20 kDa molecules of Schistosoma japonicum Chinese mainland strain. Methods Based on the inserted sequence of one of the positive clones of Schistosoma japonicum c DNA library, one pair of primers was designed and synthesized. The largest open reading frame was amplified by PCR and cloned into p GEM-T vector , And the nucleotide sequence of the cloned product was determined and analyzed. The on-line software was used to deduce the sequence of the encoded amino acid. Results A single DNA band was amplified from the inserted fragment by PCR. The size ranged from 5 15 bp to 6 97 bp. The product was cloned into p GEM-T vector. The results of double digestion and PCR showed that the fragment was identical with the previous one. The nucleotide sequence of the PCR product showed a total length of 56 bp. The online software analysis showed that the open reading frame This fragment encodes 187 amino acids and encodes a peptide with a molecular weight of about 20.2 kDa. Conclusion The experiment successfully amplified and cloned the target gene fragment, which laid the foundation for further study