After linearization, the constructed vectors of pcDNA3. 1 -hTERT and pcDNA3.1 - SV40 T were co - transfected into the Holstein bovine mammary epithelial cells to research the role of human telomer-ase reverse transcriptase (hTERT) and simian virus 40 large T transfected antigen ( SV40 T) on the culture of mammary epithelial cells in vitro. The cultured cells were detected by RT - PCR and identified by immunoflu-orescence method. The results showed that the co - expression of hTERT and SV40 T in bovine mammary epithelial cells could effectively prolong the cell culture time, increase the passage number, and the obtained cells could normally expressing keratin. In a word, the expression of hTERT and SV40 T in cultured mammary cells could effectively prolong cell life and did not influence the mammary cell characteristics.%构建pcDNA3.1-hTERT、pcDNA3.1-SV40 T载体,线性化后,共转染荷斯坦奶牛乳腺上皮细胞,研究人端粒酶逆转录酶(hTERT)和猿猴病毒40大T抗原(SV40 T)对荷斯坦奶牛乳腺上皮细胞体外培养的作用,并对细胞进行RT-PCR检测分析及免疫荧光鉴定.结果表明,hTERT和SV40 T在奶牛乳腺上皮细胞中表达可以有效地延长细胞的体外培养时间,增加细胞传代次数,获得的细胞系可以正常表达角蛋白.说明在体外培养的乳腺细胞中共表达hTERT和SV40 T可以有效延长细胞寿命且不影响乳腺细胞特性.
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