本文選題：白細(xì)胞介素 + 質(zhì)粒構(gòu)建�。� 參考：《生物技術(shù)通報(bào)》2016年02期

【摘要】：構(gòu)建人白細(xì)胞介素24(IL-24)原核表達(dá)載體并利用ELP-Intein系統(tǒng)表達(dá)純化可溶性的IL-24蛋白。通過PCR擴(kuò)增不含信號肽的人IL-24基因,將IL-24基因插入p ET-ELP-Intein質(zhì)粒構(gòu)建重組表達(dá)載體p ET-ELP-Intein-IL-24。將重組質(zhì)粒轉(zhuǎn)化至E.coliBLR(DE3),20℃下經(jīng)IPTG誘導(dǎo)表達(dá)。利用ELP蛋白在不同溫度下發(fā)生相變的特點(diǎn)和Intein蛋白的自切割反應(yīng)純化可溶性IL-24蛋白,將純化得到的IL-24蛋白進(jìn)行Western blot鑒定。用Annexin V-FITC/PI細(xì)胞凋亡檢測試劑盒檢測IL-24蛋白的生物學(xué)活性。成功構(gòu)建了重組表達(dá)載體p ET-ELP-Intein-IL-24,第一次通過原核表達(dá)方法表達(dá)并純化出了可溶性的IL-24蛋白。Western blot檢測顯示目的蛋白能與IL-24抗體特異性結(jié)合,表明純化出的蛋白確實(shí)為IL-24蛋白。細(xì)胞凋亡檢測實(shí)驗(yàn)證明IL-24蛋白能顯著地誘導(dǎo)hep G2細(xì)胞發(fā)生凋亡。
[Abstract]:The prokaryotic expression vector of human interleukin-24 (IL-24) was constructed and the soluble IL-24 protein was expressed and purified by ELP-Intein system.The human IL-24 gene without signal peptide was amplified by PCR, and the IL-24 gene was inserted into p ET-ELP-Intein plasmid to construct the recombinant expression vector pET-ELP-Intein-IL-24.The recombinant plasmid was transformed into E. coli BLRD3 and expressed at 20 鈩，

本文編號：1768845