【摘要】 将ＰＣＲ法分别从绵羊和人血基因组ＤＮＡ中扩增出的绵羊β－乳球蛋白基因（ＢＬＧ）５′端调控区８９８ｂｐ的片段和人骨髓单核细胞表面分化抗原ＣＤ１４基因（ｈＣＤ１４）成熟肽１２４５ｂｐ的片段连接．连接片段插入ｐＵＣ１９ＥｃｏＲＩ－ＫｐｎＩ位点，构建成ｐＢＬＧ－ｈＣＤ１４质粒．该质粒经ＫｐｎＩ及ＥｃｏＲＩ－ＨｉｎｄＩＩ酶切、ＰＣＲ法扩增ＢＬＧ和ｈＣＤ１４分析后，进一步用３２Ｐ－ＢＬＧ探针杂交确证．用ＥｃｏＲＩ－ＨｉｎｄＩＩ酶切质粒片段ＢＬＧ－ｈＣＤ１４产生转基因小鼠以检测ＢＬＧ－ｈＣＤ１４在乳腺的表达能力．用免疫杂交法证实在转基因阳性小鼠乳汁中表达了ｈＣＤ１４．这一结果表明ＰＣＲ法构建转基因动物特异性表达基因是可行的更多还原

【Abstract】 A 898 bp 5 ′ flanking DNA of the sheep β lactoglobulin gene (BLG) and a 1245 bp encoding region of the human myelomonocytic surface differentiation antigen CD14 gene (hCD14), from PCR using the genomic DNA of sheep and human as template respectively, were inserted into Eco RI KpnI site of pUC19 to construct pBLG hCD14. After identificated by KpnI and Eco RI HindIII restriction analyses, the pBLG hCD14 was hybridized with 32 P labelled BLG 898 bp probe. The positive clone was digested by Eco RI HindIII. The BLG hCD14 fragment was microinjected into mouse pronucleus to produce transgenic mouse. Immunoblotting was performed to investigate the expression of BLG hCD14 in transgenic mouse milk.In the present study, hCD14 was expressed by transgenic mouse mammary gland. The results showed that it is available for constructing non milk protein fusion gene to express in mammary gland using PCR.更多还原