爪蟾
ATP结合盒运输机
生物
克隆(编程)
运输机
细胞生物学
计算生物学
生物化学
基因
计算机科学
程序设计语言
作者
Deyang Xu,Dorottya Veres,Zeinu Mussa Belew,Carl Erik Olsen,Hussam Hassan Nour-Eldin,Barbara Ann Halkier
出处
期刊:Methods in Enzymology
日期:2016-01-01
卷期号:: 207-224
被引量:10
标识
DOI:10.1016/bs.mie.2016.04.001
摘要
Transport engineering in bioengineering is aimed at efficient export of the final product to reduce toxicity and feedback inhibition and to increase yield. The ATP-binding cassette (ABC) transporters with their highly diverse substrate specificity and role in cellular efflux are potentially suitable in transport engineering approaches, although their size and high number of introns make them notoriously difficult to clone. Here, we report a novel in planta "exon engineering" strategy for cloning of full-length coding sequence of ABC transporters followed by methods for biochemical characterization of ABC exporters in Xenopus oocytes. Although the Xenopus oocyte expression system is particularly suitable for expression of membrane proteins and powerful in screening for novel transporter activity, only few examples of successful expression of ABC transporter has been reported. This raises the question whether the oocytes system is suitable to express and characterize ABC transporters. Thus we have selected AtABCG25, previously characterized in insect cells as the exporter of commercially valuable abscisic acid-as case study for optimizing of characterization in Xenopus oocytes. The tools provided will hopefully contribute to more successful transport engineering in synthetic biology.
科研通智能强力驱动
Strongly Powered by AbleSci AI