An Improved Single-Step Cloning Strategy Simplifies the Agrobacterium tumefaciens-Mediated Transformation (ATMT)-Based Gene-Disruption Method for Verticillium dahliae

Phytopathology
Sheng WangJie Zhang

Abstract

The soilborne fungal pathogen Verticillium dahliae infects a broad range of plant species to cause severe diseases. The availability of Verticillium genome sequences has provided opportunities for large-scale investigations of individual gene function in Verticillium strains using Agrobacterium tumefaciens-mediated transformation (ATMT)-based gene-disruption strategies. Traditional ATMT vectors require multiple cloning steps and elaborate characterization procedures to achieve successful gene replacement; thus, these vectors are not suitable for high-throughput ATMT-based gene deletion. Several advancements have been made that either involve simplification of the steps required for gene-deletion vector construction or increase the efficiency of the technique for rapid recombinant characterization. However, an ATMT binary vector that is both simple and efficient is still lacking. Here, we generated a USER-ATMT dual-selection (DS) binary vector, which combines both the advantages of the USER single-step cloning technique and the efficiency of the herpes simplex virus thymidine kinase negative-selection marker. Highly efficient deletion of three different genes in V. dahliae using the USER-ATMT-DS vector enabled verification that ...Continue Reading

References

Aug 6, 2004·Applied and Environmental Microbiology·Jian-Ying WangXiao-Ya Chen
May 17, 2005·Fungal Genetics and Biology : FG & B·Chang Hyun KhangSeogchan Kang
Feb 6, 2008·Fungal Genetics and Biology : FG & B·María D García-PedrajasScott E Gold
Apr 24, 2009·Annual Review of Phytopathology·Steven J KlostermanKrishna V Subbarao
Dec 5, 2009·Molecular Plant-microbe Interactions : MPMI·Jun YangYou-Liang Peng
Dec 21, 2011·Methods in Molecular Biology·Rasmus John Normand FrandsenHenriette Giese
Sep 14, 2012·Molecular Plant-microbe Interactions : MPMI·Parthasarathy Santhanam, Bart P H J Thomma

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Citations

Mar 16, 2017·Molecular Plant-microbe Interactions : MPMI·Chengjian XieXingyong Yang
Mar 27, 2018·Environmental Microbiology·Zhidi FengZhaosheng Kong
Apr 11, 2019·Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences·Yun JinHui-Shan Guo
Aug 3, 2021·Frontiers in Microbiology·Jorge Luis Sarmiento-VillamilLouis Bernier

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