中国畜牧兽医2017，Vol.44Issue(11)：3130-3136,7.DOI:10.16431/j.cnki.1671-7236.2017.11.003
猪β2肾上腺素能受体基因真核表达载体及突变体的构建及表达
Construction and Expression of the Eukaryotic Expression Plasmid and Mutant of β2 Adrenergic Receptor of Pig
摘要
Abstract
This study was aimed to construct the eukaryotic expression plasmid of β2 adrenergic receptor (β2 AR) and mutants including D130N,C285S and D130N/C285S and demonstrate the plasmids could be expressed in HEK293T successfully.The β2 AR gene was amplified by PCR from pig genome DNA,then subcloned into pcDNA3.1 (+) vector by homologous recombination to construct the eukaryotic expression plasmid,added the c-myc tag in the plasmid which named myc-pcDNA3.1(+)-β2 AR.We made the eukaryotic expression vector as the template to construct the mutant,transfected the plasmids into HEK293T,extracted the protein and demonstrated the expression by Western blotting.The results of sequencing showed that the 130th amino acid had been mutate from aspartic acid to asparagine,the 285th amino acid had been mutate from cysteine to serine.Also,we confirmed that the plasmids could be expressed in HEK293T successfully.The results showed that the recombinant plasmid myc-pcDNA3.1 (+)-β2 AR,mycpcDNA3.1(+)-β2 AR-D130N,myc-pcDNA3.1 (+)-β2 AR-C285S and myc-pcDNA3.1 (+)-β2 AR-D130N/C285S were constructed correctly and could be expressed in HEK293T,which laid the foundation for further studies on the function and the pharmacology activity of β2 AR.关键词
猪/β2 AR基因/真核表达载体/突变体/Western blottingKey words
pig/β2AR gene/eukaryotic expression vector/mutant/Western blotting分类
农业科技引用本文复制引用
张海洁,谢华杰,江礼捷,连俊伟,刘敏,闵天奇,王志强..猪β2肾上腺素能受体基因真核表达载体及突变体的构建及表达[J].中国畜牧兽医,2017,44(11):3130-3136,7.基金项目
江苏高校优势学科建设工程项目(PAPD) （PAPD）
