SB转座子介导的hLF乳腺表达载体的构建及鉴定
SB전좌자개도적hLF유선표체재체적구건급감정
Construction and identification of mammary gland expression vector of human lactoferrin mediated by Sleeping Beauty transposon
  • 万方数据
    江苏农业学报 강소농업학보
    JIANGSU JOURNAL OF AGRICULTURAL SCIENCES
    2011年 4期 795-801 ,共7页

    宋洋%王子玉%张艳丽%钟部帅%张国敏%孟立%黄荣%王昌龙%闫益波%王锋

    송양%왕자옥%장염려%종부수%장국민%맹립%황영%왕창룡%염익파%왕봉

    “睡美人”(SB)转座子%β-乳球蛋白%乳腺表达载体%乳腺上皮细胞 “睡美人”(SB)轉座子%β-乳毬蛋白%乳腺錶達載體%乳腺上皮細胞
    “수미인”(SB)전좌자%β-유구단백%유선표체재체%유선상피세포
    构建了以睡美人( Sleeping beauty,SB)转座子为载体元件、β-乳球蛋白质(β-lactoglobulin,BLG)为调控区的人乳铁蛋白质(Human lactoferrin,hLF)乳腺表达载体及增强型绿色荧光蛋白质(EGFP)表达载体,便于验证SB转座子的转座效率,为hLF乳腺表达载体成功转染乳腺上皮细胞(GMC)奠定基础.采用PCR法从绵羊基因组中扩增5′、3′BLG,经TA克隆,在中间载体pcDNA3.1(-)的作用下,与hLF连接,得到pcDNA3.1(-)-BLG-hLF,然后与pT2连接,构建pT2-BLG-hLF；此外,还构建了pT2-BLG-EGFP.采用测序、酶切、PCR 3种方法鉴定阳性克隆,并用脂质体法将pT2-BLG-EGFP转染至GMC细胞中进行观察.分子鉴定结果:5′BLG、3′BLG克隆成功,pT2-BIG-hLF及pT2-BLG-EGFP构建正确；细胞鉴定结果:SB转座子介导的EGFP能在GMC细胞中高效表达.成功克隆了5′BLG、3′BLG调控区,并成功构建了乳腺表达载体pT2-BLG-hLF.
    구건료이수미인( Sleeping beauty,SB)전좌자위재체원건、β-유구단백질(β-lactoglobulin,BLG)위조공구적인유철단백질(Human lactoferrin,hLF)유선표체재체급증강형록색형광단백질(EGFP)표체재체,편우험증SB전좌자적전좌효솔,위hLF유선표체재체성공전염유선상피세포(GMC)전정기출.채용PCR법종면양기인조중확증5′、3′BLG,경TA극륭,재중간재체pcDNA3.1(-)적작용하,여hLF련접,득도pcDNA3.1(-)-BLG-hLF,연후여pT2련접,구건pT2-BLG-hLF；차외,환구건료pT2-BLG-EGFP.채용측서、매절、PCR 3충방법감정양성극륭,병용지질체법장pT2-BLG-EGFP전염지GMC세포중진행관찰.분자감정결과:5′BLG、3′BLG극륭성공,pT2-BIG-hLF급pT2-BLG-EGFP구건정학；세포감정결과:SB전좌자개도적EGFP능재GMC세포중고효표체.성공극륭료5′BLG、3′BLG조공구,병성공구건료유선표체재체pT2-BLG-hLF.
    원문보기 원문다운로드 사이트로이동
저자의 최근 논문