期刊
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
卷 97, 期 1, 页码 283-295出版社
SPRINGER
DOI: 10.1007/s00253-012-4561-7
关键词
Basidiomycete; Rhodotorula slooffiae; Rhodosporidium kratochvilovae; beta-Carotene; RNAi; T-DNA
资金
- United States National Science Foundation [MCB-0920581]
- Italian Ministry of Education, University and Scientific Research [2008JKH2MM]
- Italian Ministry of Foreign Affairs [LS-7]
- Department of Agricultural, Environmental and Food Sciences, University of Molise, Italy
The red yeasts of the Pucciniomycotina have rarely been transformed with DNA molecules. Transformation methods were recently developed for a species of Sporobolomyces, based on selection using uracil auxotrophs and plasmids carrying the wild-type copies of the URA3 and URA5 genes. However, these plasmids were ineffective in the transformation of closely related species. Using the genome-sequenced strain of Rhodotorula graminis as a starting point, the URA3 and URA5 genes were cloned and tested for the transformation ability into different Pucciniomycotina species by biolistic and Agrobacterium-mediated transformations. Transformation success depended on the red yeast species and the origin of the URA3 or URA5 genes, which may be related to the high G + C DNA content found in several species. A new vector was generated to confer resistance to nourseothricin, using a native promoter from R. graminis and the naturally high G + C nourseothricin acetyltransferease gene. This provides a second selectable marker in these species. Targeted gene disruption was tested in Sporobolomyces sp. IAM 13481 using different lengths of homologous DNA with biolistic and Agrobacterium transformation methods. Both DNA delivery methods were effective for targeted replacement of a gene required for carotenoid pigment biosynthesis. The constructs also triggered transgene silencing. These developments open the way to identify and manipulate gene functions in a large group of basidiomycete fungi.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据