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当前位置: 首页 > 文章 > pCDNA3-PTH真核表达载体构建及其在细胞内表达 畜牧与兽医 2008,40 (10) 1-4
Position: Home > Articles > Construction of eukaryotic expressing plasmid of chicken PTH gene and its expression in cells Animal Husbandry & Veterinary Medicine 2008,40 (10) 1-4

pCDNA3-PTH真核表达载体构建及其在细胞内表达

作  者：
俞路;章世元;王雅倩;刘波;李治学;周联高;严桂芹;汪益峰;林显华
单  位：
扬州大学动物科学与技术学院
关键词：
甲状旁腺素;转染;间接免疫荧光试验(IFA)
摘  要：
以鸡甲状旁腺组织中总RNA为模板,采用RT-PCR法获得甲状旁腺素(PTH)cDNA。将该基因克隆至pCDNA3载体中,构建真核细胞表达载体pCDNA3-PTH,DNA测序证明获得了PTH基因,其序列与GenBank报道序列比较,核苷酸同源性为99.4%,在第159位处存在C→A的有意义突变。将测序正确的PTH基因在多聚乙肽亚胺(PEI)介导下转染COS-1细胞,通过间接免疫荧光试验(IFA)检测到了PTH在COS-1中的表达。PTH基因克隆和表达均获得成功,为进一步探讨该基因在改善蛋壳质量中的应用奠定了基础。
译  名：
Construction of eukaryotic expressing plasmid of chicken PTH gene and its expression in cells
作  者：
YU Lu,ZHANG Shi-yuan,WANG Ya-qian,LIU Bo,LI Zhi-xue,ZHOU Lian-gao,YAN Gui-qin,WANG Yi-feng,LIN Xian-hua(College of Animal Science and Technology,Yangzhou University,Yangzhou 225009,China)
关键词：
parathyroid hormone;transfection;IFA
摘  要：
Total RNA was extracted from chicken parathyroid gland tissue,and then the parathyroid hormone(PTH) cDNA was obtained by RT-PCR.The PTH gene fragment was cloned into pCDNA3 vector,and thus the pCDNA3-PTH vector was constructed.The PTH gene encoding 119 amino acids was 360 bp in length,and the homologous comparison showed 99.4% with the sequence reported in GenBank(M36522.1),except that one nucleotide changed from C to A at 159 base site.The recombinant plasmid was transfected into COS-1 cells.At 72 hours after transfection,the expression of PTH was confirmed by indirect immunofluorescence assay(IFA) with the antiserum of PTH expressed in COS-1.This study laid a foundation for further applications of PTH in diagnosis and improving eggshell quality.

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