蓝氏贾第鞭毛虫表面变异抗原基因的克隆与序列测定(英文)

Cloning and Sequencing of the Gene Encoding Variant-Specific Surface Antigen from Giardia lamblia

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 sequence variant-specific surface antigen gene from Giardia lamblia isolate SUCH/89/BTMRI/2（C2） derived from human in China. Methods Total genomic DNA of G. lamblia was extracted and a full-length variant-specific surface antigen gene fragment was amplified by polymerase chain reaction （PCR）. The PCR product was cloned into pMD19-T simple-vector, transformed into an Escherichia coli JM109 strain and then sequenced. The sequence analysis for cloned fragment was finished by Vector NTI 9.0 software for the homology of Giardia variantspecific surface antigen gene to that of sequences publishend in GenBank. Results The full-length variant-specific surface antigen gene fragment from G. lamblia was found to be 2 142 bp, encoding a 713 amino acid polypeptide and contained a single open reading frame （ORF）. The deduced polypeptide sequence was rich in cysteine （11.8 mol%）, most of which occurred with in 29 copies of the 4-amino acid CXXC motif, one GGCY-tetrapeptide motifs and three NXS consensus N-linked glycosylation sites. This polypeptide was also rich in threonine （10.2 mol%）, glycine （12.1 mol%） and alanine （10.1 mol%）. Like other previously identified VSPs, it contained a highly conserved hydrophobic Cterminal region. The homology of G. lamblia SUCH/89/BTMRI/2（C2） variant-specific surface antigen gene to that of sequence （TSA417） published in GenBank was 99% both at the nueleotide and the amino acid levels. Conclusion The full length variant-specific surface antigen gene from the isolate of G. lamblia has the common characteristics with other previously identified VSPs.