摘要
目的 筛选日本血吸虫 (Schistosomajaponicum ,Sj)新的疫苗候选基因或雌虫性别特异性基因。 方法 用Sj雌虫抗原免疫家兔制备血清 ,对Sj成虫cDNA文库进行免疫筛选 ,将阳性克隆的插入片段进行PCR扩增 ,采用Sanger双脱氧核苷酸末端终止法对阳性克隆插入子进行测序 ,与GeneBank中的已知序列进行比对分析。 结果 共获得 2 6个阳性克隆 ,其PCR产物大小约 0 .5～ 3kb。在进行测序的 8个阳性克隆中发现了GeneBank未报道的Sj肌球蛋白的部分基因序列 (GeneBank登录号为AY3 2 2 14 8)和 1个新基因 (命名为Sj F1,GeneBank登录号为AY2 61995 )。Sj F1编码的蛋白理论分子质量为 13 .45ku ,等电点为 6.2 9,富含α螺旋和随机卷曲 ,含多个蛋白激酶磷酸化位点 ,1个酪氨酸激酶Ⅱ磷酸化位点和 1个豆蔻酸位点。 结论 Sj雌虫免疫血清可识别Sj特异性抗原分子 ,该抗原基因是否为雌虫性别特异性基因及其作为日本血吸虫病疫苗候选基因的价值 ,有待进一步研究。
Objective To find out new effective Schistosoma japonicum vaccine candidate genes or female worm sex-specific genes.Methods S. japonicum adult worm cDNA library was screened with sera from rabbits immunized with S. japonicum female worm antigen. The inserts of positive clones were identified firstly by PCR and further by sequencing and data analysis through internet Nucleotide BLAST software of NCBI and Expert Protein Analysis System of Swiss Institute of Bioinformatics. Results The 26 positive clones were obtained after re-screening. Their PCR product sizes ranged from 0.5～3 kb. A new partial length gene of S. japonicum myosin and another new gene (designated Sj-F1) were discovered and registered in GeneBank (GeneBank accession number are AY322148 and AY261995). The protein has a deduced molecular mass of 13.45 ku and isoelectric point of 6.29. Sj-F1 protein was rich in α-helix and random coil,and contained several protein kinase phosphorylation sites,one casein kinase Ⅱ phosphorylation site and one N-myristoylation site. Conclusion The sera from rabbits immunized with S. japonicum female worm could recognize S. japonicum specific antigens. It was worthwhile to further study the immune protection of Sj-F1,and further investigation on whether it is a female worm gender-specific gene is of great attraction.
出处
《中国寄生虫病防治杂志》
CSCD
2004年第1期38-41,共4页
Chinese Journal of Parasitic Disease Control
基金
TDR/WHO资助项目 (No .980 2 68)
关键词
血吸虫
日本
雌虫
CDNA文库
免疫筛选
序列分析
Schistosoma japonicum
female worm
cDNA library
immunoscreening
sequence analysis
