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目的克隆并测定中国人源蓝氏贾第鞭毛虫滋养体SUCH/89/BTMRI/2(C2)的表面变异抗原基因序列。方法提取蓝氏贾第鞭毛虫基因组DNA,PCR扩增表面变异抗原基因片段。将PCR产物连接到pMD19-T载体, 并转化至大肠埃希菌JM109中进行序列测定。通过NTI9.0软件对该基因的核苷酸和氨基酸序列进行分析,同时与基因库中已发表的蓝氏贾第鞭毛虫表面变异抗原基因进行同源序列比对。结果中国人源蓝氏贾第鞭毛虫基因全长为2142bp,编码1条长为713个氨基酸残基的多肽链,含有一个简单的开放阅读框 (ORF)。这一推测的多肽链序列富含半胱氨酸(11.8 mol%),且以CXXC模基序重复出现29次、GGCY四肽模基序出现1次及NXS重复3次在N端连接糖基化位点中。此外,这条多肽链还富含苏氨酸 (10.2 mol%) 、甘氨酸(12.1mol%)和丙氨酸(10.1 mol%)。同其他已确定的表面变异抗原(VSPs)一样,中国人源蓝氏贾第鞭毛虫 SUCH/89/BTMRI/2 (C2) 株的表面变异抗原也含有高度保守的疏水性 C 末端。该表面变异抗原基因的核苷酸序列及推导的氨基酸序列同GenBank中Gillin发表的TSA417序列的同源性均高达 99%。结论中国人源蓝氏贾第鞭毛虫滋养体表达的表面变异抗原基因具有与已鉴定的蓝氏贾第鞭毛虫表面变异抗原基因同样的特性。
Objective To clone and determine the gene sequence of surface variant antigen of Chinese human Lymanstardia gondii tropism SUCH / 89 / BTMRI / 2 (C2). Methods The genomic DNA of Giardia lamblia was extracted and the gene fragment of surface variant antigen was amplified by PCR. The PCR product was ligated into the pMD19-T vector and transformed into Escherichia coli JM109 for sequence determination. The NTI9.0 software was used to analyze the nucleotide and amino acid sequence of the gene and the homology sequences of the published gene of surface antigen of Giardia lamblia in the gene bank. Results The full-length gene of Chinese human Giardia lamblia was 2142bp and encoded a polypeptide chain of 713 amino acid residues with a simple open reading frame (ORF). This deduced polypeptide chain sequence is rich in cysteine (11.8 mol%) and appears repeatedly 29 times with the CXXC motif, the GGCY tetrapeptide motif appears once and the NXS repeats three times at the N-terminus In the locus. In addition, this polypeptide chain is also rich in threonine (10.2 mol%), glycine (12.1 mol%) and alanine (10.1 mol%). Like other established surface-variant antigens (VSPs), the surface-variant antigens of the Chinese human Giardia lamblia SCP / 89 / BTMRI / 2 (C2) strain also contain a highly conserved hydrophobic C-terminus. The nucleotide sequence and deduced amino acid sequence of the surface variant antigen gene are up to 99% homologous to the TSA417 sequence published by Gillin in GenBank. CONCLUSION: The gene of surface variant antigen expressed by Chinese human Giardia lamblia trophozoites has the same characteristics as the identified gene of surface antigen of Giardia lamblia.