人血清白蛋白载体的构建及其在山羊乳腺中的瞬时表达

根据GeneBank中人血清白蛋白(HSA)的基因序列,设计一对特异性的引物,从人肝cDNA文库中扩增出其编码序列,并加入酶切位点XhoⅠ,以pCDNA 3.1为模板,设计特异性的引物,并在上游引物加入NotⅠ,下游加入ApaⅠ和NotⅠ,扩增出筛选标记新霉素抗性基因neo,然后再以pBC1为基本骨架,在其NotⅠ、XhoⅠ位点处分别插入hsa和neo,从而得到乳腺表达载体pBC-NEO-HSA,将该质粒注射入奶山羊乳腺中进行暂时性的表达,结果显示所构建的载体能够有效地指导目的基因的表达,从而为下一步构建细胞系、克隆动物作好基础。

Construction of human serum albumin gland-specific expression vector and its transient expression in goat mammary glands

According to the gene sequence of the human serum albumin in GeneBank, a pair of specific primers, which was introduced a enzyme site XhoⅠ at the 5′-end, were designed. Screening tag neo was amplified by PCR from the pCDNA 3.1 vector using a pair of specific primers which was added two enzyme site NotⅠ and ApaⅠ. The mammary gland-specific expression vector pBC-NEO-HSA was constructed by inserting the neo and hsa into the enzyme site NotⅠ and XhoⅠ in the pBC1 vector, respectively. The pBC-NEO-HSA vector was injected directly into the lactating mammary glands of goats. The transient expression results indicated that the mammary specific expression vector driven by the goat β-casein gene promoter could efficiently direct the expression of human serum albumin in goat milk. This study provides the basis to get the established cell line and further study on the production of the transgenic animal.