Journal
JOURNAL OF MICROBIOLOGICAL METHODS
Volume 84, Issue 3, Pages 418-422Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.mimet.2011.01.017
Keywords
Agrobacterium tumefaciens; Dimorphic fungus; Sporothrix schenckii; T-DNA flanking sequences; Transformation
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Funding
- National Natural Science Foundation of China [30910103903, 81071334]
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Sporothrix schenckii is a dimorphic pathogenic fungus that causes human and animal sporotrichosis globally. Here we developed and optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) system of S. schenckii for insertional mutagenesis. The transformation efficiency reached more than 600 transformants per 10(6) conidia. Using this protocol enabled us to obtain a large number of T-DNA insertional mutants within a short experimental period. Several mutants with altered phenotypes were obtained during the transformation experiments. The mutants displayed mitotic stability. Transferred DNA (T-DNA) flanking sequences were cloned by thermal asymmetric interlaced PCR (TAIL-PCR). Our results demonstrated that the ATMT system can be an effective tool for insertional mutagenesis in S. schenckii. This is the first report of a suitable mutagenesis system which may provide valuable mutants and information for both forward and reverse genetics research in the future for this medically important fungus. (C) 2011 Elsevier B.V. All rights reserved.
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