资源描述：
《新城疫病毒mukteswar株f基因重组pgapz_f的构建》由会员上传分享，免费在线阅读，更多相关内容在教育资源-天天文库。
1、第31卷第6期江西农业大学学报Vol.31,No.62009年12月ActaAgriculturaeUniversitatisJiangxiensisDec.,2009文章编号:1000-2286(2009)06-1079-04新城疫病毒Mukteswar株F基因重组pGAPZα-F的构建121程太平,荣俊,刘超(1.长江大学动物科学学院,湖北荆州434025;2.长江大学生命科学学院,湖北荆州434025)摘要:以RT-PCR扩增新城疫病毒Mukteswar株F基因F1片段和F2片段,以核酸内切酶KpnⅠ和Xb
2、aⅠ对目的基因片段及质粒pGAPZαA进行酶切,连接酶切产物,转化E.coliDH5α。以PCR方法确定MukteswarF1和MukteswarF2的阳性重组子均为4个。对阳性重组子进行酶切鉴定及序列分析,结果,重组质粒pGAPZαA-F1、pGAPZαA-F2及质粒pGAPZαA的酶切电泳条带与试验设计大小相符;基因测序得到的重组子中F1和F2序列长度分别为1198bp、269bp,与新城疫病毒Mukteswar株F基因序列比对,其序列长度和核苷酸排列完全一致。结果表明重组质粒中目的基因片段的核苷酸序列、大小
3、和插入位置是正确的,为以酵母表达系统表达F1和F2,研究Mukteswar株与基因Ⅶ型毒株之间F蛋白的抗原性差异程度打下基础。关键词:新城疫病毒Mukteswar株;F基因;质粒pGAPZαA;重组中图分类号:S852.65文献标识码:ARecombinationofPlasmidpGAPZαandFGeneofNewcastleDiseaseVirusStrainMukteswar121CHENGTai-ping,RONGJun,LIUChao(1.CollegeofAnimalScience,YangtzeU
4、niversity,Jingzhou434025,China;2.CollegeofLifeScience,YangtzeUniversity,Jingzhou434025,China)Abstract:TheF1andF2ofFgeneofNewcastlediseasevirusstrainMukteswarwereamplifiedwithre2versetranscription-polymerasechainreaction,thegoalgenepiecesandplasmidpGAPZαAwered
5、igestedwithKpnⅠandXbaⅠ,thedigestedpieceswerelinkedwithDNAligase,E.coliDH5αweretransformedbythelinkedoutcomes.Positiverecombinantsweredistinguishedwithpolymerasechainreaction,andpositivere2combinantsofMukteswarF1andMukteswarF2were4colonieseach.Thepositiverecom
6、binantswereidentifiedwithendonucleasedigestingandDNAsequenceanalysis.RecombinantplasmidpGAPZαA-F1,pGAPZαA-F2andplasmidpGAPZαAweredigestedwithKpnⅠandXbaⅠ,thepositionoftheirbandsinagarosegelafterelectrophoresiscorrespondedtothetestdesign.F1sequenceandF2sequence
7、wererespectively1198bpand269bpinrecombinantplasmidpGAPZαA-F1andpGAPZαA-F2,theirsequencesshowednodifferenceingenelengthandnucleotidesequenceswithFgeneofNewcastlediseasevirusisolateMukteswarbycontrasting.Theseresultsindicatethatthenucleotidesequence,size,insert
8、edsiteofthefragmentofF1andF2arecorrectinrecombinantplasmidpGAPZαA-F1andpGAPZαA-F2,asthebasisforexpressingF1andF2with收稿日期:2009-06-09修回日期:2009-09-16基金项目:湖北省教育厅科学研究计划项目(D200612005)作者简介:程太平(1
