A kit and method for rapidly detecting chemical endocrine disrupting activity
A technology of endocrine disruption and chemicals, applied in biochemical equipment and methods, biological testing, material inspection products, etc., can solve the problems of less toxic targets, low throughput, and cumbersome operation in one test
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[0044] 1. Prokaryotic expression and purification of human estrogen-related receptor γ-ligand binding domain hERRγ-LBD recombinant protein
[0045] The total RNA of human colorectal adenocarcinoma cell Caco-2 was extracted with Trizol reagent, and reversed into cDNA as a template. Referring to the coding sequence of human estrogen-related receptor ERRγ (NCBI GenBank: NP_001230435.1) published by NCBI, primers were designed to amplify its sequence. Using the double enzyme digestion method, the coding gene of hERRγ-LBD (44..263aa) was connected to the pGEX-4T-1 vector, transformed into E.coli DH5α competent cells, and the positive clones were screened and sequenced to obtain the recombinant expression vector pGEX-4T - hERRγ-LBD. The recombinant plasmid was extracted, transformed into E.coli BL21 competent cells, and hERRγ-LBD recombinant expression bacteria were obtained. The recombinant bacteria were amplified and cultured at 37°C, and the expression was induced by adding IPT...
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