Abstract: Methods are provided for converting into a sequence specific strand specific and location specific DNA nicking endonuclease, a restriction endonuclease that recognizes an asymmetric DNA sequence, the endonuclease having two catalytic sites and one or more single sequence specific DNA-binding domains. In one embodiment the method requires inactivating one of the catalytic sites of the restriction endonuclease. In another embodiment, the restriction endonuclease is a dimer having a first and second subunit each comprising a sequence specific DNA binding domain, a catalytic site and a dimerization domain. The nicking endonuclease is formed from combining one subunit having an inactivated catalytic site and a second subunit having an inactivated DNA binding domain. The nicking endonuclease may be converted into a restriction endonuclease by the addition of manganese cations in the digestion buffer.

Abstract: The present invention relates to: recombinant DNA encoding the SbfI restriction endonuclease as well as the SbfI methylase, and expression of the SbfI restriction endonuclease and SbfI methylase in E. coli cells containing the recombinant DNA; and methods for cloning the SbfI restriction gene (sbfIR) from Streptomyces species Bf-61 into E. coli by PCR. The method relied on primers based on DNA sequences predicted from amino acid sequences of the purified SbfI restriction endonuclease.

Abstract: The present invention relates to recombinant DNA which encodes a novel nicking endonuclease, N.BstNBI, and the production of N.BstNBI restriction endonuclease from the recombinant DNA utilizing PleI modification methylase. Related expression vectors, as well as the application of N.BstNBI and other nicking enzymes in non-modified strand displacement amplification, is disclosed also.

Abstract: Methods are provided for converting into a sequence specific strand specific and location specific DNA nicking endonuclease, a restriction endonuclease that recognizes an asymmetric DNA sequence, the endonuclease having two catalytic sites and one or more single sequence specific DNA-binding domains. In one embodiment the method requires inactivating one of the catalytic sites of the restriction endonuclease. In another embodiment, the restriction endonuclease is a dimer having a first and second subunit each comprising a sequence specific DNA binding domain, a catalytic site and a dimerization domain. The nicking endonuclease is formed from combining one subunit having an inactivated catalytic site and a second subunit having an inactivated DNA binding domain. The nicking endonuclease may be converted into a restriction endonuclease by the addition of manganese cations in the digestion buffer.

Abstract: The present invention relates to recombinant DNA which encodes the BstYI restriction endonuclease as well as BstYI methyltransferase, expression of BstYI restriction endonuclease and M.BstYI in E. coli cells containing the recombinant DNA. It also relates to methods for purification of the recombinant BstYI restriction endonuclease and BstYI methyltransferase.

Abstract: RsaI, a restriction enzyme from the bacterium Rhodopseudomonas sphaeroides, recognizes the DNA sequence 5′-GTAC-3′. Because RsaI is commercially valuable, we sought to overproduce it by cloning the genes for RsaI and its accompanying, modification, enzyme. The ‘methylase-selection’ method, the customary procedure for cloning restriction and modification genes, was applied to RsaI. The method yielded clones containing the methylase gene (rsaIM), but none containing both the methylase gene and the restriction gene (rsaIR). Inverse-PCR was then used to recover sections of the DNA downstream of rsaIM. These sections were sequenced, and the sequences were joined in silico to reveal the gene organization of the RsaI R-M system.

Abstract: The methylase selection method was used to clone the SnaBI methylase gene (snaBIM) from Sphaerotilus natans (ATCC 15291). An active SnaBI methylase was cloned in E. coli using pSnaBI-2, a pUC19 derivative containing two SnaBI sites. Because methylase and restriction genes are usually located alongside each other in a restriction-modification systems, efforts were made to clone the downstream DNA by inverse PCR. Inverse PCR cloned the missing portion of the SnaBI endonuclease and also identified a control, or C, protein. The two open reading frames snaBIR (ORF1) and snaBIC (ORF2) converged toward the SnaBI methylase gene (ORF). Attempts to establish a snaBIR-recombinant plasmid expressing the SnaBI endonuclease in E.coli modified with SnaBI methylase failed. Overexpression of the SnaBI endonuclease in E. coli required the use of the heterospecific BsaAI methylase.

Abstract: A recombinant restriction endonuclease from Anabaena variabilis is provided, as well as the isolated gene which encodes it and methods for the production of the recombinant AvaI restriction endonuclease. An isolated gene encoding a modification methylase from A. variabilis is also provided.

Abstract: In accordance with the present invention there is provided an isolated DNA coding for the Bg/I restriction endonuclease and modification methylase derived from Bacillus globigii RUB561 stain, as well as related methods for cloning said recombinant DNA. The present invention also relates to clones which express recombinant Bg/I restriction endonuclease and recombinant modification methylase produced from Bg/I recombinant DNA and to methods for producing said enzymes.

Abstract: The present invention provides a novel approach to the production of restriction enzymes. More specifically, there is provided a novel method for cloning these enzymes, which comprises preparing DNA libraries from the DNA of an organism that synthesizes the restriction enzyme of interest, creating a suitable host containing a heterospecific methyltransferase gene able to protectively modify DNA from digestion by the restriction enzyme of interest, introducing the DNA libraries into the protectively modified host, and screening recombinant organisms to identify those carrying the desired restriction enzyme gene.The application of this method to the FspI and HaeIII restriction genes of Fischerella species and Haemophilus aegyptius, respectively, is described in detail, together with the resulting clones that form the basis of a new and useful process for purifying the FspI and HaeIII restriction enzymes.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to digestion by the restriction enzyme and subsequent selection of clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: Methods for cloning restriction enzymes and their corresponding modification enzymes by selecting clones resistant to in vitro digestion by the restriction enzyme and subsequent screening to identify those clones containing the restriction gene.

Abstract: The present invention provides a novel approach to the production of restriction enzymes. More specifically, there is provided a novel method for cloning these enzymes, which comprises preparing DNA libraries from the DNA of an organism that synthesizes the restriction enzyme of interest, creating a suitable host containing a heterospecific methyltransferase gene able to protectively modify DNA from digestion by the restriction enzyme of interest, introducing the DNA libraries into the protectively modified host, and screening recombinant organisms to identify those carrying the desired restriction enzyme gene.The application of this method to the FspI and HaeIII restriction genes of Fischerella species and Haemophilus aegyptius, respectively, is described in detail, together with the resulting clones that form the basis of a new and useful process for purifying the FspI and HaeIII restriction enzymes.

Abstract: The present invention is directed to a method for cloning and producing the MwoI restriction endonuclease by 1) introducing the restriction endonuclease gene from M. wolfei into a host whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid encoding and expressing the MwoI restriction endonuclease activity, and 3) purifying the MwoI restriction endonuclease from the fermented host which contains the plasmid encoding and expressing the MwoI restriction endonuclease activity.

Abstract: The present invention is directed to a method for cloning and producing the Afl II restriction endonuclease by 1) introducing the restriction endonuclease gene from Anabaena flos-aquae into a host whereby the restriction gene is expressed; 2) fermenting the host which contains the plasmid encoding and expressing the Afl II restriction endonuclease activity, and 3) purifying the Afl II restriction endonuclease from the fermented host which contains the plasmid encoding and expressing the Afl II restriction endonuclease activity.