中国寄生虫学与寄生虫病杂志 ›› 2009, Vol. 27 ›› Issue (3): 20-228.

• 研究简报 • 上一篇    下一篇

马来丝虫3-磷酸甘油醛脱氢酶基因的克隆、序列分析及编码产物B细胞表位预测

谢东方, 方政*, 童海燕, 徐邦生, 黄为群, 方浩, 沈勤   

  1. 南通大学医学院寄生虫学教研室, 南通 226001
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2009-06-30 发布日期:2009-06-30
  • 通讯作者: 方政

Cloning, Sequencing of G3PD Gene from Brugia malayi and Prediction of B cell Epitopes in its Amino Acid Sequence

XIE Dong-fang, FANG Zheng*, TONG Hai-yan, XU Bang-sheng, HUANG Wei-qun, FANG Hao, SHEN Qin   

  1. Department of Parasitology, School of Basic Medical Sciences, Nantong University, Nantong 226001, China
  • Received:1900-01-01 Revised:1900-01-01 Online:2009-06-30 Published:2009-06-30
  • Contact: FANG Zheng

摘要: 根据GenBank中马来丝虫3-磷酸甘油醛脱氢酶基因(BmG3PD基因)序列设计引物, 以马来丝虫mRNA为模板, RT-PCR扩增BmG3PD基因, 将其克隆入pGEM-T载体, 转化大肠埃希菌(E. coli)DH5α, 筛选阳性克隆。经EcoRⅠ和XhoⅠ双酶切及PCR鉴定, 获得阳性重组质粒pGEM-BmG3PD, 经序列分析及同源性比较, 以及对其编码产物进行B细胞表位预测, 结果表明PCR扩增的特异性条带为1 020 bp, 与预期相符, 与GenBank已知基因序列同源性为99%。编码产物B细胞表位预测, 氨基酸区域可能在22~36、242~255、303~318和326~336位。

关键词: 马来丝虫, 3-磷酸甘油醛脱氢酶, 基因克隆, 序列分析, B细胞表位

Abstract:

Specific primers were designed and synthesized based on the

reported glyceraldehydes-3-phosphate dehydrogenase(BmG3PD) gene of Brugia malayi(GenBank Accession No. U18137). Total RNA was extracted from

Brugia malayi and its BmG3PD gene was amplified

by reverse transcription-polymerase chain raction(RT-PCR). The PCR product was purified and cloned into plasmid pGEM-T, then transformed into Escherichia coli DH5α. The recombinant

plasmids were screened and identified by digestion

with restriction enzyme and PCR amplification. The positive recombinant plasmid pGEM-T-BmG3PD was confirmed by sequencing and homology comparison. Five parameters and

methods were used to predict B-cell epitopes in amino acid sequence of BmG3PD. The amplified DNA fragment(1 020 bp) had a high identity of 99% with the BmG3PD gene sequence of

Brugia malayi. B-cell epitopes of BmG3PD were probably at or adjacent to 22-36, 242-255, 303-318 and 326-336 in its amino acid sequence.

Key words: Brugia malayi, G3PD, Gene cloning, Sequence analysis, B-cell epitope