摘要
目的构建乙型肝炎病毒(HBV)全基因组C区L97和V60突变质粒,并检测其生物学活性。方法采用定点突变技术将HBVadr1.2拷贝基因组质粒p3.8Ⅱ进行C区点突变,构建L97和V60两个突变株。质粒经脂质体介导转染HepG2细胞,Southern杂交分析细胞内HBVDNA复制,ELISA检测培养上清中HBsAg和HBeAg。结果测序结果表明所构建的突变质粒p3.8L97和p3.8V60分别为nt2189A→C和nt2086C→G,突变质粒均可在宿主细胞内复制和表达。结论所构建HBV(adr型)全基因组C区L97和V60突变质粒具有生物活性。
Objective To constructthecoremutant(L97andV60)plasmidsof hepatitisB virus(HBV)andaassesstheir biologicalactivity.Methods Site-directedmutagenesiswas performedto inducespecificcorepointmutationsin HBVadr suhtype 1.2copygenomeplasmidp3.8Ⅱ.TheplasmidsweretransfectedintoHepG2cellsvia liposom,andintracellularHBV DNAwas analyzedby Southernhybridization,whileextracellularHBVantigens(HBsAgand HBeAg)in the culture supernatantwereassayed by enzyme-linkedimmunosorbentassay.Results Sequenceanalysisof the constructedmutant plasmidsp3.8L97andp3.8V60demonstratedmutationsat2189A→C and2086C→G respectively.Thesemutantplasmids exhibitedHBVDNAreplicationactivityandgeneexpressioninhostcells.Conclusion HBVadrsubtypegenomecoremutant(L97andV60)plasmidswe haveconstructedpossessbiologicalactivity.
出处
《第一军医大学学报》
CSCD
北大核心
2002年第3期254-255,261,共3页
Journal of First Military Medical University
