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Optimal conditions for genetic transformation of the cyanobacterium Anacystis nidulans R2
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Under optimal conditions, the cyanobacterium Anacystis nidulans R2 was transformed to ampicillin resistance at frequencies of greater than 10(7) transformants per microgram of plasmid (pCH1) donor DNA. No stringent period of competency was detected, and high frequencies of transformation were achieved with cultures at various growth stages. Transformation increased with time after addition of donor DNA up to 15 to 18 h. The peak of transformation efficiency (transformants/donor molecule) occurred at plasmid concentrations of 125 to 325 ng/ml with an ampicillin resistance donor plasmid (pCH1) and 300 to 625 ng/ml for chloramphenicol resistance conferred by plasmid pSG111. The efficiency of transformation was enhanced by excluding light during the incubation or by blocking photosynthesis with the electron transport inhibitor 3-(3, 4-dichlorophenyl)-1, 1-dimethylurea (DCMU) or the uncoupler carbonyl cyanide-m-chlorophenyl hydrazone. Preincubation of cells in darkness for 15 to 18 h before addition of donor DNA significantly decreased transformation efficiency. Growth of cells in iron-deficient medium before transformation enhanced efficiency fourfold. These results were obtained with selection for ampicillin (pCH1 donor plasmid)- or chloramphenicol (pSG111 donor plasmid)-resistant transformants. Approximately 1,000 transformants per microgram were obtained when chromosomal DNA from an herbicide (DCMU)-resistant mutant was used as donor DNA. DCMU resistance was also transferred to recipient cells by using restriction fragments of chromosomal DNA from DCMU-resistant mutants. This procedure allowed size classes of fragments to be assayed for the presence of the DCMU resistance gene.
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Metadata
- Authors
- Golden, S. S., & Sherman, L. A.
- Venue
- Journal of Bacteriology
- Publisher
- American Society for Microbiology
- Published
- 1984-04
- Volume
- 158
- Issue
- 1
- Pages
- 36-42
- ISSN
- 0021-9193 (ISSN-L)
- 1098-5530
- PMC
- PMC215375
- OpenAlex
- W1687442175
- Cited by
- 174
- References
- 19
- Citations by year
- 2026: 4 · 2025: 2 · 2024: 2 · 2023: 11 · 2022: 2 · 2021: 11 · 2020: 2 · 2019: 1 · 2018: 6 · 2017: 1 · 2016: 4 · 2015: 5 · 2014: 7 · 2013: 3 · 2012: 5
- FWCI
- 4.50
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Journal of Bacteriology (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Photosynthetic Processes and Mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
- Keywords
- Anacystis nidulans, cyanobacteria, genetic transformation, transformation efficiency, ampicillin resistance, chloramphenicol resistance, photosynthesis inhibition, iron deficiency, restriction fragment mapping, resistance gene transfer
- MeSH terms
- Cyanobacteria, Ampicillin, Binding Sites, Carbonyl Cyanide m-Chlorophenyl Hydrazone, Chloramphenicol, Culture Media, Diuron, DNA, Genes, Iron, Kinetics, Penicillin Resistance, Photosynthesis, Transformation, Genetic
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