重组腺病毒介导的人凝血Ⅸ因子cDNA在奶山羊乳腺中的分泌表达

Human Clotting Factor IX(hFIX)Secretion in Goat Milk afterTransfer with hFIX cDNA into Mammary Gland by Using Recombinant Adenovirus Vector

利用含ｈＦＩＸｃＤＮＡ和腺病毒片段的质粒ｐＡｄＣＭＶｈＦＩＸ与质粒ｐＪＭ１７共转染２９３包装细胞，制备含ｈＦＩＸ的重组腺病毒颗粒，经纯化、浓缩后通过直接转染活体奶山羊乳腺小叶的方法，建立了ｈＦＩＸ蛋白的乳腺表达系统。转染处理４天后，ｈＦＩＸ蛋白在羊奶中的最高表达量为２１ｎｇ／ｍｌ乳汁，活性检测表明乳汁中的人凝血ＩＸ因子蛋白８０％以上具有生物学活性。此研究结果证实了人凝血ＩＸ因子全长ｃＤＮＡ在奶山羊乳汁中分泌表达的可行性，同时也为验证外源基因在奶山羊乳腺组织中的分泌表达提供了一个快速检测系统。

Adenovirus particles carrying human clotting factor IX (hFIX) cDNA were produced after co- transfecting 293 packaging cells with plasmid pAdCMVhFIX and pJM17. An in vivo hFIX ex- pression system in goat mammary gland was constructed by transfecting of recombinant adeovirus into mammary gland lobule of lactating goats. The highest production of hFIX in goat milk was 21ng/ml 4 days after transfection. Activity analysis showed that >80% hFIX protein in milk was bilologically active. These results confirm that hFIX cDNA could express in goat mammary gland , and also provide a rapid detective system for evaluation of a foreign gene expression and secreation in milk.