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3-酮基嘌呤还原酶(TSC10)基因表达载体的构建及其在毕赤酵母中的表达

Construction of 3-keto Purine Reductase (TSC10) Gene Expression Vector and Its Expression in Pichia Pastoris
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摘要 TSC10基因所编码的3-酮基嘌呤还原酶是酵母中神经酰胺合成的重要因子。设计了一种从酵母中提取神经酰胺经济、便捷、高效的方法:(1)利用构建含有TSC10基因的毕赤酵母GS115表达载体p PIC3.5K-TSC10;(2)电转化法将该表达质粒转化到GS115感受态细胞中;(3)用G418筛选以及PCR鉴定;(4)使用qRT-PCR和SDS-PAGE进行检测。结果经过G418筛选以及PCR鉴定后确定获得了包含TSC10基因的毕赤酵母转化子,经过qRT-PCR和SDS-PAGE进行检测后发现TSC10基因在20个阳性菌株中均可以稳定、高效表达。成功构建了3-酮基嘌呤还原酶高表达的毕赤酵母菌株,并为后续获得高收率神经酰胺奠定了基础。 The 3-keto purine reductase encoded by TSC10 gene is an important factor in the synthesis of ceramide in yeast.The economical,convenient and efficient method was designed for extracting ceramide from yeast.①Using the expression vector pPIC3.5K-TSC10 containing the TSC10 gene of Pichia pastoris GS115.②The expression plasmid was transformed into GS115 competent cells by electroporation.③Screened with G418 and identified by PCR.④Detection was performed using qRT-PCR and SDS-PAGE.After purification by G418 and PCR identification,the Pichia pastoris transformants containing TSC10 gene were identified and analyzed by qRT-PCR and SDS-PAGE.The TSC10 gene was stable and highly expressed in 20 positive strains.This method successfully constructed the highly expressed Pichia pastoris of 3-ketopurine reductase and laid the foundation for the subsequent high yield of ceramide.
作者 郑丽娜 刘美辰 王思明 白雪媛 赵雨 赵大庆 王佳雯 ZHENG Li-na;LIU Mei-chen;WANG Si-ming;BAI Xue-yuan;ZHAO Yu;ZHAO Da-qing;WANG Jia-wen(Traditional Chinese Medicine and Biotechnology Research and Development Center,Changchun University of Chinese Medicine,Changchun 130117,China)
出处 《科学技术与工程》 北大核心 2018年第10期31-34,共4页 Science Technology and Engineering
基金 吉林省中药组学工程实验室建设项目(2014N155)资助
关键词 3-酮基嘌呤还原酶 毕赤酵母 基因表达 3-ketopurine reductase Pichia pastoris gene expression
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  • 1何学民.神经酰胺——一种新型高级化妆品生物活性添加剂[J].中国化妆品(专业版),1994(9):30-31. 被引量:1
  • 2为什么说米糠是个宝(2)[J].农产品加工,2005(4):28-29. 被引量:2
  • 3刘创,陈成良.当代化妆品中的几种热门原料概述[J].黑龙江日化,1996(1):36-37. 被引量:3
  • 4熊曼萍.神经酰胺的美容作用.中国保健食品,2005,5(1):12-12.
  • 5[1]Goodman Y,Mattson MP.Ceramide protects hippocampalneurons against excitotoxic and oxidative insults,and amyloidbeta-peptide toxicity[J].J Neurochem,1996,66(2):869-872.
  • 6[2]Hannun Y A.Functions of ceramide in coordinating cellularresponses to stress[J].Science,1996,274(13):1855.
  • 7[3]Merrill A H.,J.A.Shayman,R.T.Riley.Sphingolipids-the enigmatic lipid class:Biochemistry,physiology,and pathophysiology[J].Toxicology and Applied Pharmacology,1997,142:208.
  • 8[4]Schmelz E M,Merrill Jr.Sphingomyelin consumption sup-presses aberrant colonic crypt foci and increases the proportion of adenomas versus adenocarcinomas in CF1 mice treated with 1,2-dimethylhydrazine:Implications for dietary sphingolipids and colon carcinogenesis[J].Cancer Research,1996,56(1):4936.
  • 9[5]Spiegel S,A.H.Merrill Jr.Sphingolipid metabolism and cell growth regulation[J].FASEB,1996,10:1388.
  • 10[6]Chmura S J,Loss of ceramide production confers resistance to radiation-induced apoptosis[J].Cancer Research,1997,57(1):1270.

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