摘要
构建HBV C基因型YIDD拉米夫定耐药株的全基因真核表达载体,为进一步深入探讨乙型肝炎病毒拉米夫定耐药的分子机制,寻求耐药后的有效治疗奠定基础。以重组质粒PMD18T-HBV-C为模板,采用PCR扩增HBV C基因型YIDD变异株的全基因组DNA,并将其定向克隆于真核表达载体PcDNA3.1(+)中。获得的真核表达重组质粒PcDNA3.1(+)-HBV-C(YIDD)通过酶切后电泳及测序进行鉴定。琼脂糖电泳结果证实PCR产物大小约为3.2 kb,与预期相同。酶切及测序结果证实,HBV C基因型YIDD拉米夫定耐药株全基因真核表达载体PcDNA3.1(+)-HBV-C(YIDD)构建成功。
This study aimed to construct a eukaryotic recombinant expression vector containing entire C genotype （YIDD） genome of HBV. PCR was performed to amplify the full genomic DNA of variant YIDD belonging to genotype C of HBV by using recombinant plasmid PMD18T-HBV as the template. A new recombinant plasmid named as PcDNA3, 1 （ ＋ ）-HBV-C（YIDD） was constructed through inserting the genomic DNA into PcDNA3.1 （ ＋ ）. This eukaryotic expression vector was identified by restriction endonuclease map and sequencing analysis, The result of agarose electrophoresis showed a 3, 2 kb PCR product in size that was the same as the expected. The results of endonuclease digestion and sequencing confirmed that a eukaryotic expression vector PcDNA3.1 （ ＋ ） -HBV-C （YIDD） containing entire genomic DNA of variant YIDD belonging to HBV genotype C was successfully constructed.
出处
《微生物学杂志》
CAS
CSCD
2007年第5期39-42,共4页
Journal of Microbiology
基金
高校博士点专项基金项目(20050226002)
哈尔滨医科大学研究生创新基金
关键词
乙型肝炎病毒拉米夫定耐药变异真核表达载体
Hepatitis B virus
Lamivudine drug-resistant mutation
Eukaryotic expression vector
