Feng Xin-Gang, Lin Jiao-Jiao, Zhu Jian-Guo, Zhang Liang, Yuan Chun-Xiu, Cai You-Min
Shanghai Institute of Animal Parasitology, Chinese Academy of Agricultural Sciences, Key Laboratory of Animal Parasitology, Ministry of Agriculture of China, Shanghai 200232, China.
Sheng Wu Gong Cheng Xue Bao. 2002 Jan;18(2):231-4.
The cDNA fragments of interest were amplified using Sj lambda ZipLox library as the templates by PCR and then cloned into a eukaryotic expression vector p-CMV-GH; A small number of DNA fragments inserted in the recombinants was identified by restriction cleavage, EST sequencing and bioinformatical analysis; mice were injected intramuscularly with the expression library (L-CMV-SjR) or sublibraries(L-CMV-SjR1, L-CMV-SjR2 and L-CMV-SjR3), immunized mice were challenged with Schistosoma japonicum cercariae on day 35, the levels of IgG antibodies in sera from the immunized mice were detected by ELISA. The results demonstrated that a partial cDNA expression library of S.j, with approximately 10(5) transformants, was constructed, most of the recombinants contained the insert DNA fragments of interest, and these fragments had the features of protein-coding sequences for Schistosome. There were no significant differences for the levels of IgG antibodies in sera from all of the immunized groups. Mice immunized with L-CMV-SjR, L-CMV-SjR1 and L-CMV-SjR2 developed significant protective effect against Sj infection compared to control mice injected with the empty plasmid, the rate of worm reduction was about 30%.
以日本血吸虫λ-ZipLox文库为模板,通过PCR扩增出感兴趣的cDNA片段,然后克隆到真核表达载体p-CMV-GH中;通过酶切、EST测序和生物信息学分析鉴定重组体中插入的少量DNA片段;将表达文库(L-CMV-SjR)或亚文库(L-CMV-SjR1、L-CMV-SjR2和L-CMV-SjR3)肌肉注射给小鼠,在第35天用日本血吸虫尾蚴攻击免疫小鼠,通过ELISA检测免疫小鼠血清中IgG抗体水平。结果表明,构建了一个约含10^5个转化子的日本血吸虫部分cDNA表达文库,大多数重组体含有感兴趣的插入DNA片段,这些片段具有血吸虫蛋白质编码序列的特征。所有免疫组小鼠血清中IgG抗体水平无显著差异。与注射空质粒的对照小鼠相比,用L-CMV-SjR、L-CMV-SjR1和L-CMV-SjR2免疫的小鼠对日本血吸虫感染产生了显著的保护作用,减虫率约为30%。