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US5965415A Expired / Lapsed US ⏱️ 0.0 years remaining

Process for the in vivo recombination of DNA sequences having mismatched bases

Assignee: Mixis France SA
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Filing: 1995-06-07
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Granted: 1999-10-12
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Expires: 2015-06-07
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Abstract

Abstract The present invention relates to a process of recombination in vivo of partially homologous DNA sequences having up to 30% of base mismatches. According to its essential characteristic, said sequences are placed together in cells or an organism of which the enzymatic mismatch repair system is defective or has been transitorily inactivated by saturation for the time to obtain recombination between said DNA sequences or in using mutants which increase the intergeneric recombination.

Inventors

  • 👤 Miroslav Radman
  • 👤 Christiane Rayssiguier

Bibliographic Details

Application Number —
Jurisdiction US
Data Source google_patents
Last Synchronized 2026-11-04 05:34
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/10; C12N15/102; C12N15/1027

Claims Analysis

Total Claims: 6
Claim 1
Independent Claim 341 words
A process for producing a hybrid organism, comprising: combining in a cell or cells of an organism a double-stranded DNA from a first species or genus with a double stranded DNA from a second species or genus, said first and second DNAs having sequences which are partially homologous and have mismatches able to activate the enzymatic mismatch repair system of the cell when said system is functional, wherein the enzymatic mismatch repair system is defective or has been inactivated transitorily to enable stable recombination between said DNA sequences, wherein said process is selected from the group of processes consisting of: (i) fusing cells of an organism of a first species or a first genus with cells of an organism of a second species or a second genus, said cells of said organism of a second species or a second genus having a defective enzymatic mismatch repair system or having said system inactivated transitorily; (ii) crossing a unicellular organism of a first species or a first genus with a unicellular organism of a second species or a second genus, wherein at least one organism has a defective enzymatic mismatch repair system or has said system inactivated transitorily; (iii) conjugating or transducing a recipient bacterium of a first species or a first genus and a donor bacterium of a second species or a second genus, said donor bacterium having at least one DNA sequence to be transferred to said recipient bacterium, wherein at least one of said donor bacterium and said recipient bacterium includes a defective enzymatic mismatch repair system or said system is inactivated transitorily; (iv) inactivating said enzymatic mismatch repair system by at least one mutation of at least one of the mutS and mutL genes; and (v) placing into a unicellular organism defective or inactivated in the enzymatic mismatch repair system two DNA sequences, said two DNA sequences being partially homologous genes derived from two different organisms, and selecting the desired hybrid sequence, gene, or product thereof, and selecting said cell or cells of said organism containing said recombined DNA.
Claim 2
Dependent Claim 341 words
A process for producing a hybrid organism, comprising: combining in a cell or cells of an organism a double-stranded DNA from a first species or genus with a double stranded DNA from a second species or genus, said first and second DNAs having sequences which are partially homologous and have mismatches able to activate the enzymatic mismatch repair system of the cell when said system is functional, wherein the enzymatic mismatch repair system is defective or has been inactivated transitorily to enable stable recombination between said DNA sequences, wherein said process is selected from the group of processes consisting of: (i) fusing cells of an organism of a first species or a first genus with cells of an organism of a second species or a second genus, said cells of said organism of a second species or a second genus having a defective enzymatic mismatch repair system or having said system inactivated transitorily; (ii) crossing a unicellular organism of a first species or a first genus with a unicellular organism of a second species or a second genus, wherein at least one organism has a defective enzymatic mismatch repair system or has said system inactivated transitorily; (iii) conjugating or transducing a recipient bacterium of a first species or a first genus and a donor bacterium of a second species or a second genus, said donor bacterium having at least one DNA sequence to be transferred to said recipient bacterium, wherein at least one of said donor bacterium and said recipient bacterium includes a defective enzymatic mismatch repair system or said system is inactivated transitorily; (iv) inactivating said enzymatic mismatch repair system by at least one mutation of at least one of the mutS and mutL genes; and (v) placing into a unicellular organism defective or inactivated in the enzymatic mismatch repair system two DNA sequences, said two DNA sequences being partially homologous genes derived from two different organisms, and selecting the desired hybrid sequence, gene, or product thereof, and selecting said cell or cells of said organism containing said recombined DNA.
Claim 3
Dependent Claim 21 words
The process of claim 1, wherein said DNA is recombined in a chromosome of said cell or cells of said organism.
Claim 4
Dependent Claim 336 words
A process for producing a hybrid protein, comprising: combining in a cell a double-stranded DNA from a first species or genus with a double stranded DNA from a second species or genus, said first and second DNAs having sequences which are partially homologous and have mismatches able to activate the enzymatic mismatch repair system of the cell when said system is functional, wherein the enzymatic mismatch repair system is defective or has been inactivated transitorily to enable stable recombination between said DNA sequences, wherein said process is selected from the group of processes consisting of: (i) fusing cells of an organism of a first species or a first genus with cells of an organism of a second species or a second genus, said cells of said organism of a second species or a second genus having a defective enzymatic mismatch repair system or having said system inactivated transitorily; (ii) crossing a unicellular organism of a first species or a first genus with a unicellular organism of a second species or a second genus, wherein at least one organism has a defective enzymatic mismatch repair system or has said system inactivated transitorily; (iii) conjugating or transducing a recipient bacterium of a first species or a first genus and a donor bacterium of a second species or a second genus, said donor bacterium having at least one DNA sequence to be transferred to said recipient bacterium, wherein at least one of said donor bacterium and said recipient bacterium includes a defective enzymatic mismatch repair system or said system is inactivated transitorily; (iv) inactivating said enzymatic mismatch repair system by at least one mutation of at least one of the mutS and mutL genes; and (v) placing into a unicellular organism defective or inactivated in the enzymatic mismatch repair system two DNA sequences, said two DNA sequences being partially homologous genes derived from two different organisms, and selecting the desired hybrid sequence, gene, or product thereof, and expressing said recombined DNA in an organism to produce said hybrid protein.
Claim 5
Dependent Claim 336 words
A process for producing a hybrid protein, comprising: combining in a cell a double-stranded DNA from a first species or genus with a double stranded DNA from a second species or genus, said first and second DNAs having sequences which are partially homologous and have mismatches able to activate the enzymatic mismatch repair system of the cell when said system is functional, wherein the enzymatic mismatch repair system is defective or has been inactivated transitorily to enable stable recombination between said DNA sequences, wherein said process is selected from the group of processes consisting of: (i) fusing cells of an organism of a first species or a first genus with cells of an organism of a second species or a second genus, said cells of said organism of a second species or a second genus having a defective enzymatic mismatch repair system or having said system inactivated transitorily; (ii) crossing a unicellular organism of a first species or a first genus with a unicellular organism of a second species or a second genus, wherein at least one organism has a defective enzymatic mismatch repair system or has said system inactivated transitorily; (iii) conjugating or transducing a recipient bacterium of a first species or a first genus and a donor bacterium of a second species or a second genus, said donor bacterium having at least one DNA sequence to be transferred to said recipient bacterium, wherein at least one of said donor bacterium and said recipient bacterium includes a defective enzymatic mismatch repair system or said system is inactivated transitorily; (iv) inactivating said enzymatic mismatch repair system by at least one mutation of at least one of the mutS and mutL genes; and (v) placing into a unicellular organism defective or inactivated in the enzymatic mismatch repair system two DNA sequences, said two DNA sequences being partially homologous genes derived from two different organisms, and selecting the desired hybrid sequence, gene, or product thereof, and expressing said recombined DNA in an organism to produce said hybrid protein.
Claim 6
Dependent Claim 36 words
The process of part (v) of claim 3, wherein each DNA sequence is contained within a separate plasmid, wherein each plasmid is introduced into a bacterium which includes a defective or inactivated mismatched DNA repair system.

References Cited (Backward Citations)

US4863727A Cited
US4963469A Cited

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