研究了β2肾上腺素受体（β2 adrenergic receptor，β2AR）基因重组表达质粒在人胚肾细胞293（Human embryonic kidney cells 293，HEK293）中的瞬时转染及表达产物的纯化和鉴定。将已构建的重组表达质粒瞬时转染HEK293细胞，表达产物制备粗膜蛋白后，采用镍离子亲和层析法纯化得到受体蛋白并进行鉴定。结果表明：重组表达质粒pTriEx-1.1 Hygro-β2AR1-418转染HEK293细胞后表达量和蛋白活性较优，最佳表达时间为转染后72 h。SDS-PAGE和Western blot结果显示，在47 ku 左右出现了特异性条带，咪唑的最佳洗脱浓度为250 mmol/L，受体蛋白纯度大于80%。活性鉴定表明：受体蛋白与HRP酶标记的3种β激动剂均能特异性结合，检测HRP-克伦特罗、HRP-莱克多巴胺和HRP-沙丁胺醇的OD值分别为0.97、0.91和0.94。通过纯化得到的受体蛋白，约占粗膜蛋白的4.4%。本试验成功获得一定数量纯度和活性较好的β2AR受体蛋白，为受体技术在β激动剂多残留检测中的研发和应用奠定了基础。

The transient transfection of β2AR gene to mammalian cells HEK293 was investigated, and then the expressed product was purified and identified. The constructed recombinant plasmid was transiently transfected into HEK293 cells. Then the expressed product was treated by crude membrane preparation and purification through Ni-NTA-affinity chromatography. Subsequently, the content and activity of the recombinant protein was identified. The results showed that the HEK293 cells transfected with pTriEx-1.1 Hygro-β2AR1-418 exhibited higher expression effect, and the optimal expression was 72 h after transfection. The results of SDS-PAGE and Western blot indicated that a specific band was seen at 47 ku, the optimal imidazole concentration was 250 mmol/L, and the purity of the protein was more than 80%. The activitiy assay revealed that the recombinant receptor could bind all 3 HRP-β-agonists, and the OD values obtained from the ELISA for detection HRP-CBL, HRP-RAC and HRP-SAL were 0.97, 0.91 and 0.94，respectively. The yield of the receptor protein was 4.42% of crude membrane. In brief, a certain amount of β2AR protein with comparatively ideal purity and activity was obtained, laying a foundation for the development and application of β2AR in multi-residues determination of β-agonists.

国家公益性行业（农业）科研专项（201203094）