凡纳滨对虾P53亲水肽段原核表达及抗体制备
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摘要
在哺乳动物细胞中,肿瘤抑制基因P53是目前研究最为广泛和系统的抑癌基因之一。P53参与了DNA损伤修复、细胞周期调控、细胞凋亡及抑制血管生成等过程,在甲壳动物中研究较少。对Lv P53的蛋白质全序列进行抗原性分析,将编码亲水肽段DNA连接到Pet32a表达载体上,转化原核表达菌株BL21,利用异丙基-β-D-硫代半乳糖苷(isopropylβ-D-thiogalactoside,简称IPTG)诱导融合蛋白质表达,经蛋白电泳分析表明,融合蛋白主要以包涵体形式存在。利用Ni柱亲和纯化发酵产物,得到较单一的蛋白,以制备多克隆抗体。
引文
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[3]Chanock S J,Benna J,Smith R M,et al.The respiratory burst oxidase[J].Journal of Biological Chemistry,1994,269(40):24519-24522.
[4]Mu1oz M,Cede1o R,Rodríquez J,et al.Measurement of reactive Oxygen intermediate production in haemocytes of the penaeid shrimp,Penaeus vannamei[J].Aquaculture,2000(191):89-107.
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[6]Serrano M A,Li Z,Mohan D,et al.DNA-PK,ATM and ATR collaboratively regulate p53-RPA interaction to facilitate homologous recombination DNA repair[J].Oncogene,2013,32(19):2452.
[7]Pani G,Koch O R,Galeotti T.The p53-p66shc-manganese superoxide dismutase(Mn SOD)network:a mitochondrial intrigue to generate reactive oxygen species[J].International Journal of Biochemistry&Cell Biology,2009,41(5):1002-1005.
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