The DOI matches its Crossref record; 21 of the 21 references it cites were found in a registry or an index.
Oligonucleotide-directed mutagenesis using M13-derived vectors: an efficient and general procedure for the production of point mutations in any fragment of DNA
Open accessAbstract
This paper presents a versatile and efficient procedure for the construction of oligodeoxyribonucleotide directed site-specific mutations in DNA fragments cloned into M13 derived vectors. As an example, production of a transition mutation in a clone of the yeast MATa1 gene is described. The oligonucleotide is hybridized to the template DNA and covalently closed closed double stranded molecules are generated by extension of the oligonucleotide primer with E. coli DNA polymerase (large fragment) and ligation with T4 DNA ligase. The resulting double stranded closed circular DNA (CC-DNA) is separated from unligated and incompletely extended molecules by alkaline sucrose gradient centrifugation. This purification is essential for production of mutants at high efficiency. Competent E. coli JM101 cells are transformed with the CC-DNA fraction and single stranded DNA is isolated from individual plaques. The recombinants are screened for mutant molecules by 1) restriction endonuclease screening for the loss of the Hinf I site in the target region, and 2) by dot blot hybridization using the mutagenic oligonucleotide as probe. Double stranded DNA is isolated from the sequencing. Efficiency of mutant production is in the range of 10-45% and no precautions to prevent mismatch repair are required.
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Cites
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Cited by
Multiple Mutations in HIV-1 Reverse Transcriptase Confer High-Level Resistance to Zidovudine (AZT)
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Characterization of ribosomal frameshifting in HIV-1 gag-pol expression
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Comparative Anatomy of 16-S-like Ribosomal RNA
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Multiple upstream AUG codons mediate translational control of GCN4
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Metadata
- Authors
- Zoller, M. J., & Smith, M.
- Venue
- Nucleic Acids Research
- Publisher
- Oxford University Press (OUP)
- Published
- 1982
- Volume
- 10
- Issue
- 20
- Pages
- 6487-6500
- ISSN
- 0305-1048 (ISSN-L)
- 1362-4962
- OpenAlex
- W2084884863
- Cited by
- 745
- References
- 21
- FWCI
- 14.89
- Type
- journal article
- Language
- en
- Open access
- green
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- Zoller, M. J., & Smith, M. (1982). Oligonucleotide-directed mutagenesis using M13-derived vectors: an efficient and general procedure for the production of point mutations in any fragment of DNA. Nucleic Acids Research. https://doi.org/10.1093/nar/10.20.6487
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Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).
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