Abstract
RESTRICTION endonucleases recognise specific sequences in DNA, and these endonucleases, especially those which generate cohesive ends, have been widely used to clone DNA1. However, many DNAs lack sequences which are recognised by endonucleases such as EcoRI, HindIII or BamHI. A general method of overcoming this problem has been described recently2. This approach involves the synthesis of oligonucleotides sensitive to a specific endonuclease and the blunt end ligation of these molecules to the DNA to be cloned. In contrast, we sought a method which avoids the insertion of additional nucleotides into a DNA sequence, but depends on direct modification of DNA. If a DNA sequence differs in only one base pair from the recognition sequence of a restriction endonuclease, a particular change of this base pair will generate the proper sequence. Here we describe a way of generating restriction endonuclease cleavage sites by single base changes derived after in vitro methylation of single-stranded DNA.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 51 print issues and online access
$199.00 per year
only $3.90 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Roberts, R. J. Crit. Rev. Biochem. 4, 123–164 (1976).
Scheller, R. H., Dickerson, R. E., Boyer, H. W., Riggs, A. D. & Itakura, K. Science 196, 177–180 (1977).
Messing, J., Gronenborn, B., Muller-Hill, B. & Hofschneider, P. H. Proc. natn. Acad. Sci. U.S.A. 74, 3642–3646 (1977).
Dickson, R. C., Abelson, J., Barnes, W. M. & Reznikoff, W. S. Science 187, 27–35 (1975).
Messer, W. & Melchers, F. in The Lactose Operon (eds Beckwith, J. R. & Zipser, D.) 305–315 (Cold Spring Harbor Laboratory, Cold Spring Harbor, 1970).
Lawley, P. D. Mutat. Res. 23, 283–295 (1974).
Gerchman, L. L. & Lundlum, D. B. Biochim. biophys. Acta 308, 310–316 (1973).
Lawley, P. D. Sha, S. A. Biochem. J. 128, 117–132 (1972).
Kirtikar, D. M. & Goldthwait, D. A. Proc. natn. Acad. Sci. U.S.A. 71, 2022–2026 (1974).
Cohen, S. N., Chang, A. C. Y. & Hsu, L. Proc. natn. Acad. Sci. U.S.A. 69, 2110–2114 (1972).
Rigby, P. W. J., Dieckmann, M., Rhodes, C. & Berg, P. J. molec. Biol. 113, 237–251 (1977).
v.d.Hondel, C. A., Pennings, L. & Schoenmakers, J. G. G. Eur. J. Biochem. 68, 55–70 (1976).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
GRONENBORN, B., MESSING, J. Methylation of single-stranded DNA in vitro introduces new restriction endonuclease cleavage sites. Nature 272, 375–377 (1978). https://doi.org/10.1038/272375a0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/272375a0
This article is cited by
-
A novel prokaryotic vector for identification and selection of recombinants: Direct use of the vector for expression studies in E. coli
Microbial Cell Factories (2010)
-
Determination of Nucleotide Sequences in DNA
Bioscience Reports (2004)
-
The art and design of genetic screens: Escherichia coli
Nature Reviews Genetics (2003)
-
The early days of DNA sequences
Nature Medicine (2001)
-
Cloning single-stranded DNA
Molecular Biotechnology (1996)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.