Please use this identifier to cite or link to this item:
|Title:||A Simplified and efficient method for transformation and gene tagging of Ustilago maydis using frozen cells||Authors:||Ji, L.
|Issue Date:||Apr-2010||Citation:||Ji, L., Jiang, Z.-D., Liu, Y., Koh, C.M.J., Zhang, L.-H. (2010-04). A Simplified and efficient method for transformation and gene tagging of Ustilago maydis using frozen cells. Fungal Genetics and Biology 47 (4) : 279-287. ScholarBank@NUS Repository. https://doi.org/10.1016/j.fgb.2010.01.002||Abstract:||Ustilago maydis is an important model fungal organism for diverse studies. Little improvement has been made in the method for its transformation since the PEG-mediated transfection of spheroplasts that was reported more than 20 years ago. We have constructed binary T-DNA vectors carrying Hygromycin and Nourseothricin resistance gene cassettes and have developed a highly efficient method for transformation of this fungus based on Agrobacterium tumefaciens-mediated transformation (ATMT). Through a series of optimization, at least 1 × 104 Hygromycin B resistant colony forming units (CFU) have been achieved on each 90 mm agar plate using 106 sporidia. Optimal pH value for ATMT is approximately 5.6. Approximately 96% Hygromycin B-resistant transformants contain a single-copy T-DNA inserted into the nuclear genome. Analysis of 204 T-DNA flanking sequences showed that 15.2% of them were found in the coding sequences and a further 37.25% within 0.5 kb from the coding sequences at the 5′ UTR or promoter regions. In addition, a method for preparation and preservation of transformation-ready T-DNA donor and receptor cells has been developed allowing gene tagging experiments to be performed on-demand. An initial screening of 5000 mutants resulted in the identification of a putative farnesyl transferase beta subunit and a PRE6 homologue as new players of sexual mating in U. maydis. © 2010 Elsevier Inc. All rights reserved.||Source Title:||Fungal Genetics and Biology||URI:||http://scholarbank.nus.edu.sg/handle/10635/99965||ISSN:||10871845||DOI:||10.1016/j.fgb.2010.01.002|
|Appears in Collections:||Staff Publications|
Show full item record
Files in This Item:
There are no files associated with this item.
checked on Jul 10, 2020
WEB OF SCIENCETM
checked on Jul 10, 2020
checked on Jul 12, 2020
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.