CAJ | 학술논문

目的:克隆、表达与纯化出AP-2β蛋白质,并用所表达的蛋白质制备出抗AP-2β的抗体.方法:将AP-2β基因插入到表达载体pGEX-4T-1谷胱苷肽-S-转移酶基因下游,所得克隆经测序,以确定其阅读框码的正确性,然后用正确的克隆质粒转化E.coli BL21,用异丙基硫代半乳糖苷(IPTG)诱导,而后用谷胱苷肽琼脂糖-4B纯化所表达的融合蛋白质.通过凝胶迁移率变动分析实验(EMSA)测得蛋白质的生物学活性,所得蛋白质用于制备抗体.结果:得到一条大约78 KD的蛋白质,并且成功的制备了抗体,所得蛋白质和抗体均具有很好的生物学活性.结论:我们成功的表达出转录激活因子AP-2β并制备出抗AP-2β的抗体,为一下步对AP-2β的功能进行研究打下了良好的基础.
목적:극륭、표체여순화출AP-2β단백질,병용소표체적단백질제비출항AP-2β적항체.방법:장AP-2β기인삽입도표체재체pGEX-4T-1곡광감태-S-전이매기인하유,소득극륭경측서,이학정기열독광마적정학성,연후용정학적극륭질립전화E.coli BL21,용이병기류대반유당감(IPTG)유도,이후용곡광감태경지당-4B순화소표체적융합단백질.통과응효천이솔변동분석실험(EMSA)측득단백질적생물학활성,소득단백질용우제비항체.결과:득도일조대약78 KD적단백질,병차성공적제비료항체,소득단백질화항체균구유흔호적생물학활성.결론:아문성공적표체출전록격활인자AP-2β병제비출항AP-2β적항체,위일하보대AP-2β적공능진행연구타하료량호적기출.
Objective: The aim of this research is to clone,express and purify the AP-2β gene and prepare the antibody.Methods:The AP-2β gene was fused to the glutathion-S-transferase gene in the expression vector pGEX-4T-1.The clones were sequenced to confirm the reading frame. The correct clones were then used to transform E.coli BL21 and the expression of fusion protein was induced by isopropyl-β-D-thiogalaotopyranoside (IPTG). The AP-2β fusion protein was purified with the Glutathione Sepharose-4B.The activity of protein was detected by electrophoretic mobility shift assay (EMSA). Then the protein was used to immunize rabbit in order to produce anti- AP-2β serum.Results:The 78KD fusion protein was obtained and anti- AP-2β antibody was successfully prepared. The protein and the antibody have a good biological activity.Conclusions: The transcription factor AP-2β was successfully expressed and the anti- AP-2β antibody was prepared. This study provides a basis for the functional analysis of AP-2β.