Cloning and expressing restriction endonucleases and modification methylases from caryophanon
AssigneeLife Technologies Corporation
InventorsMary C. Longo, Michael D. Smith, Deb K. Chatterjee
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Abstract
The present invention is directed to recombinant hosts which contain and express the ClaI Type-II restriction endonuclease and/or modification methylase genes. The present invention is also directed to vectors or DNA molecules which contain these gene, and to methods of producing the enzymes. One source of these enzyme is Caryophanon latum, although other microorganisms may be used to isolate the restriction endonuclease isoschizomers and modification methylase isoschizomers of the invention.
Claims44
A recombinant host cell comprising a Caryophanon gene coding for a restriction endonuclease, said restriction endonuclease is capable of recognizing the palindromic sequence:
- 5'AT.dwnarw.CG AT3'
- 3'TA GC.uparw.TA5'
- and cleaving said sequence between the deoxythymidine and deoxycytidine residues, producing a two-base 5' extension.
A vector comprising a Caryophanon gene coding for a restriction endonuclease, said restriction endonuclease is capable of recognizing the palindromic sequence:
- 5'AT.dwnarw.CG AT3'
- 3'TA GC.uparw.TA5'
- and cleaving said sequence between the deoxythymidine and deoxycytidine residues, producing a two-base 5' extension.
A method of producing a restriction endonuclease which recognizes the palindromic sequence:
- 5'AT.dwnarw.CG AT3'
- 3'TA GC.uparw.TA5'
- and cleaves said sequence between the deoxythymidine and deoxycytidine residues, producing a two-base 5' extension, said method comprising:
- (a) culturing a recombinant host cell comprising a Caryophanon gene coding for said restriction endonuclease;
- and
- (b) isolating said restriction endonuclease from said host cell.
A recombinant host cell comprising a Caryophanon gene coding for a modification methylase, said modification methylase is capable of recognizing the palindromic sequence:
- 5'ATCGAT3'
- 3'TAGCTA5'
- and chemically modifying said sequence such that said modified sequence is resistant to cleavage with its corresponding restriction endonuclease.
A vector comprising a Caryophanon gene coding for a modification methylase, said modification methylase is capable of recognizing the palindromic sequence:
- 5'ATCGAT3'
- 3'TAGCTA5'
- and chemically modifying said sequence such that said modified sequence is resistant to cleavage with its corresponding restriction endonuclease.
A method of producing a modification methylase which recognizes the palindromic sequence:
- 5'ATCGAT3'
- 3'TAGCTA5'
- and modifies said sequence such that said modified sequence is resistant to cleavage with its corresponding restriction endonuclease, said method comprising:
- (a) culturing a recombinant host cell comprising a Caryophanon gene coding for said modification methylase;
- and
- (b) isolating said modification methylase from said host cell.
Dependencies are inferred from the claim text.