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High-Throughput Gene Replacementin Aspergillus fumigatus

机译：高通量基因Replacementin曲霉属真菌来自烟

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摘要

Aspergillus fumigatus is a human pathogen and the principal etiologic agentof invasive and chronic aspergillosis leading to several hundreds of thousandsof deaths every year. Very few antifungals are available to treat infectionscaused by A. fumigatus, and resistance is developing to those we have. Ourunderstanding of the molecular mechanisms that drive pathogenicity and drugresistance have been hampered by the lack of large mutant collections, whichlimits our ability to perform functional genomics analysis. Here we present ahigh-throughput gene knockout method that combines a highly reproduciblefusion PCR method to enable generation of gene replacement cassettes witha multiwell format transformation procedure. This process can be used togenerate 96 null mutants within 5 days by a single person at a cost of less than￡18 ($24) per mutant and is being employed in our laboratory to generate abarcoded genome-wide knockout library in A. fumigatus.
机译：来自烟曲霉属真菌是一种人类病原体和主要病原学的agentof入侵和慢性曲霉病导致数百名每年数千人死亡。抗真菌治疗infectionscaused a .来自烟和阻力是我们开发的。的分子机制致病性和drugresistance由于缺乏大型突变集合,whichlimits我们执行功能的能力基因组学分析。ahigh-throughput基因敲除方法结合一个高度reproduciblefusion PCR方法使一代的基因替换磁带和多井格式转换过程。零突变体由一个人在5天内成本低于￡每突变和18(24美元)被使用在我们的实验室来生成abarcoded图书馆在全基因组基因敲除。来自烟。

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  • 来源
    《Current Protocols in Microbiology》 |2019年第1期|e88/1-e88/18|共18页
  • 作者

    Can Zhao; Marcin G. Fraczek; Lauren Dineen;

  • 作者单位

    Manchester Fungal Infection Group, Division of Infection, Immunity and RespiratoryMedicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, CoreTechnology Facility, University of Manchester, Manchester, United Kingdom;

    Division of Evolution & Genomic Sciences, School of Biological Sciences, Faculty ofBiology Medicine and Health, Manchester Institute of Biotechnology, University ofManchester, Manchester, United Kingdom;

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  • 原文格式 PDF
  • 正文语种 英语
  • 中图分类
  • 关键词

    Aspergillus fumigatus; Libraries; Single Persongene replacementMutantsanimal pathogens;

    机译：来自烟曲霉属真菌;图书馆;单一PersongenereplacementMutantsanimal病原体;

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