SacB-SacR gene cassette as the negative selection marker to suppress Agrobacterium overgrowth in Agrobacterium-mediated plant transformation
Autor: | Bingyu Zhao, Yi Liu, Danyu Kong, Sy M. Traore, Jiamin Miao, Zachary L. Nimchuk, Zongrang Liu, Yiming Liu, Xunzhong Zhang |
---|---|
Jazyk: | angličtina |
Rok vydání: | 2016 |
Předmět: |
0106 biological sciences
0301 basic medicine Agrobacterium Plant tissue culture Mutant Nicotiana benthamiana Biology 01 natural sciences Biochemistry Genetics and Molecular Biology (miscellaneous) Biochemistry 03 medical and health sciences Molecular Biosciences Technology Report Molecular Biology plant transformation lcsh:QH301-705.5 2. Zero hunger Genetics SacB-SacR gene cassette fungi food and beverages Agrobacterium tumefaciens biochemical phenomena metabolism and nutrition biology.organism_classification WRKY protein domain Cell biology 030104 developmental biology Gene cassette lcsh:Biology (General) Agrobacterium overgrowth bacteria CRISPR-Cas9 010606 plant biology & botany Transformation efficiency |
Zdroj: | Frontiers in Molecular Biosciences, Vol 3 (2016) Frontiers in Molecular Biosciences |
Popis: | Agrobacterium overgrowth is a common problem in Agrobacterium-mediated plant transfor-mation. To suppress the Agrobacterium overgrowth, various antibiotics have been used during plant tissue culture steps. The antibiotics are expensive and may adversely affect plant cell differentiation and reduce plant transformation efficiency. The SacB-SacR proteins are toxic to most Agrobacterium tumefaciens strains when they are grown on culture medium sup¬plemented with sucrose. Therefore, SacB-SacR genes can be used as negative selection markers to suppress the overgrowth of Agrobacterium tumefaciens in the plant tissue culture process. We generated a mutant Agrobacterium tumefaciens strain GV2260 (recA-SacB/R) that has the SacB-SacR cassette inserted into the bacterial genome at the recA gene locus. The mutant Agrobacterium strain is sensitive to sucrose but maintains its ability to transform plant cells in both transient and stable transformation assays. We demonstrated that the mutant strain GV2260 (recA-SacB/R) can be inhibited by sucrose that reduces the overgrowth of Agrobacterium and therefore improves the plant transformation efficiency. We employed GV2260 (recA-SacB/R) to generate stable transgenic N. benthamiana plants expressing a CRISPR-Cas9 for knocking out a WRKY transcrip¬tion factor. |
Databáze: | OpenAIRE |
Externí odkaz: |