Construction and expression of a eukaryotic β<sub>3</sub>-adrenoceptor expression vector pcDNA3.1(+)-β<sub>3</sub>-AR

曾凡新; 董志; 傅洁民; 刘晓海

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Construction and expression of a eukaryotic β₃-adrenoceptor expression vector pcDNA3.1(+)-β₃-AR

更新时间：2026-01-09

- Construction and expression of a eukaryotic β₃-adrenoceptor expression vector pcDNA3.1(+)-β₃-AR
- Journal of Army Medical University Vol. 29, Issue 21, Pages: 2070-2072(2007)
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  CLC： R394-33;R394.3;R965.2
- Published：2007
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曾凡新,董志,傅洁民,刘晓海.人β3肾上腺素受体真核表达质粒pcDNA3.1(+)-β3-AR的构建及表达[J].陆军军医大学学报(原第三军医大学学报),2007,29(21):2070-2072.

　ZENG Fan-xin,DONG Zhi,FU Jie-min,LIU Xiao-hai.Construction and expression of a eukaryotic β3-adrenoceptor expression vector pcDNA3.1(+)-β3-AR[J].J Amry Med Univ (J Third Mil Med Univ),2007,29(21):2070-2072.
曾凡新,董志,傅洁民,刘晓海.人β3肾上腺素受体真核表达质粒pcDNA3.1(+)-β3-AR的构建及表达[J].陆军军医大学学报(原第三军医大学学报),2007,29(21):2070-2072. DOI：

　ZENG Fan-xin,DONG Zhi,FU Jie-min,LIU Xiao-hai.Construction and expression of a eukaryotic β3-adrenoceptor expression vector pcDNA3.1(+)-β3-AR[J].J Amry Med Univ (J Third Mil Med Univ),2007,29(21):2070-2072. DOI：

摘要

目的构建人β

肾上腺素受体（β

-AR)的真核表达质粒pcDNA3.1(+)-β

-AR，并通过脂质体介导转染HEK293细胞使之表达该受体蛋白。方法以pENTR_ADRB3-Stop质粒为模板，采用PCR法扩增出人β

-AR cDNA，将其插入真核表达载体pcDNA3.1(+)中，构建成真核表达质粒pcDNA3.1(+)-β

-AR。将该质粒转染HEK293真核细胞，通过RT-PCR检测β

-AR mRNA在HEK293细胞内的表达，通过免疫荧光法检测β

-AR蛋白在HEK293细胞中的表达。结果酶切和DNA测序鉴定均证实重组质粒含有人β

-AR编码区全长。在转染重组质粒pcDNA3.1(+)-β

-AR的HEK293细胞中检测到了β

-AR基因的转录和翻译产物。结论通过基因工程方法成功构建了β

-AR的真核表达质粒pcDNA3.1(+)-β

-AR，为进一步建立β

-AR的稳定表达细胞株，用于筛选高效、高选择性β

-AR激动剂奠定了基础。

Abstract

Objective To construct the recombinant eukaryotic expression vector pcDNA3.1(+)-β

-AR and express it in HEK293 cells. Methods Human β

-AR cDNA was amplified by PCR from a pENTR_ADRB3-Stop plasmid

and the eukaryotic expression plasmid pcDNA3.1(+)-β

-AR was constructed by inserting the β

-AR cDNA into KpnⅠ/XbaⅠ-digested pcDNA3.1(+). The DNA sequence was confirmed

by double digestion

and the pcDNA3.1(+)-β

-AR plasmid was transfected into HEK293 cell line. Human β

-AR mRNA expression on HEK293 cells was detected by RT-PCR

and the protein expression on HEK293 cells was detected by immunofluorescence technique. Results Restriction analysis and DNA sequencing showed that the recombinant plasmid contained the whole coding region of human β

-AR gene. RT-PCR assay indicated the transcription of β

-AR mRNA and the β

-AR protein detectable in the transfected HEK293 cells. Conclusion Recombinant eukaryotic expression vector pcDNA3.1(+)-β

-AR plasmid was successfully constructed by genetic engineering technique and expressed on the surface of HEK293 cells

which will contribute to further studies on the establishment of stable expression cell lines for screening high potent and selective β

-AR agonists.

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Construction and expression of a eukaryotic β3-adrenoceptor expression vector pcDNA3.1(+)-β3-AR

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Construction and expression of a eukaryotic β₃-adrenoceptor expression vector pcDNA3.1(+)-β₃-AR