Cloning of two type II methylase genes that recognise asymmetric nucleotide sequences: FokI and HgaI.
The modification genes of Flavobacterium okeanokoites and Haemophilus galinarum have been cloned into the vector pBR322 and expressed in Escherichia coli cells. FokI methylase gene is contained on a 3.80 kb piece of F. okeanokoites DNA. Plasmid constructs carrying this fragment of DNA are resistant to digestion by FokI restriction endonuclease but are sensitive to cleavage by HindIII, EcoRI and PstI. Unmodified lambda DNA molecules, exposed in vitro to cell extracts prepared from cells habouring this plasmid, became resistant to digestion by FokI. The smallest HgaI methylase clone carries the pBR322 plasmid containing a 3.50 kb piece of H. galinarum DNA. This plasmid is resistant to digestion by HgaI. Neither the FokI nor the HgaI restriction endonuclease was detected in either clone. This is the first report of cloning modification genes whose protein products recognise asymmetric nucleotide sequences.
Nwankwo
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New England Biolaboratories Inc, Beverly, MA 01915.
Wilson
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D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes / Q000235:genetics*; D015257:DNA-Cytosine Methylases; D015252:Deoxyribonucleases, Type II Site-Specific; D004926:Escherichia coli / Q000235:genetics; D005417:Flavobacterium / Q000201:enzymology / Q000235:genetics*; D005796:Genes; D005798:Genes, Bacterial; D006190:Haemophilus / Q000201:enzymology / Q000235:genetics*; D008780:Methyltransferases / Q000235:genetics*
C054689:DNA modification methylase HgaI; D015257:DNA-Cytosine Methylases; D008780:Methyltransferases; D004262:DNA Restriction Enzymes; C035042:endodeoxyribonuclease FokI; D015252:Deoxyribonucleases, Type II Site-Specific