本文選題：陰陽虛證 + 糖皮質激素受體　； 參考：《第二軍醫(yī)大學》2005年碩士論文

【摘要】：目的:構建hGRα重組原核表達質粒并進行誘導表達,為下一步研制hGRα ELISA檢測試劑盒和進一步研究GR與中醫(yī)學陰陽虛證的關系服務。 方法:以含有hGRα cDNA的重組克隆質粒pRShGRα為模板,通過PCR擴增得到hGRα的兩種基因片段,將這兩種基因片段分別克隆到pGEM-T載體上,進而亞克隆到pGEX-4T-3表達載體上。將這兩種重組表達質粒進行酶切、測序鑒定后,分別轉化大腸桿菌DH5α和BL21(DE3),并在一定的條件下進行重組蛋白的誘導表達,誘導表達產(chǎn)物行SDS-PAGE和Western blotting分析。 結果:酶切和測序結果證明hGRα的兩種基因片段均成功地重組入pGEX-4T-3載體。誘導表達產(chǎn)物的SDS-PAGE和Western blotting分析均提示有hGRα目的蛋白特異表達。 結論:本實驗成功構建了hGRα的兩種重組原核表達質粒,并成功誘導了它們的原核表達,為下一階段的研究奠定了基礎。
[Abstract]:Objective: to construct the recombinant prokaryotic expression plasmid of hGR 偽 and express it in order to further study the relationship between gr and yin and yang deficiency syndrome in traditional Chinese medicine. Methods: using the recombinant clone plasmid pRShGR 偽 containing hGR 偽 cDNA as template, two kinds of hGR 偽 gene fragments were amplified by PCR and cloned into pGEM-T vector respectively, and then subcloned into pGEX-4T-3 expression vector. The two recombinant expression plasmids were digested by enzyme and identified by sequencing. The recombinant proteins were transformed into Escherichia coli DH5 偽 and BL21DE-3, respectively. The recombinant protein was induced and expressed under certain conditions. The induced expression products were analyzed by SDS-PAGE and Western blotting. Results: the results of restriction endonuclease digestion and sequencing showed that the two gene fragments of hGR 偽 were successfully recombined into pGEX-4T-3 vector. Both SDS-PAGE and Western blotting analysis of induced expression products indicated that hGR 偽 target protein was specifically expressed. Conclusion: in this experiment, two recombinant prokaryotic expression plasmids of hGR 偽 were successfully constructed, and their prokaryotic expression was induced successfully, which laid a foundation for the next stage of research.
【學位授予單位】：第二軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2005
【分類號】：R346

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