US4637980A
Expired / Lapsed
US
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Externalization of products of bacteria
Assignee: SmithKline Beecham Corp
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Filing: 1983-08-09
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Granted: 1987-01-20
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Expires: 2003-08-09
Abstract
Abstract A bacterial product is made by transforming a temperature sensitive lysogen with a DNA molecule which codes, directly or indirectly, for the product, culturing the transformant under permissive conditions and externalizing the product by raising the temperature to induce phage encoded functions.
Inventors
- 👤 Jeffrey I. Auerbach
- 👤 Martin Rosenberg
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-01 08:32
CPC Classifications
C; C12; C12N; C12N15/00; C12N9/00; C12N9/14; C12N9/24; C12N9/2402; C12N9/2462; A; A61; A61K; A61K9/00; A61K9/48; A61K9/50; A61K9/5005; A61K9/5063; A61K9/5068; C12N1/00; C12N1/06; C12N15/09; C12N15/63; C12N15/67; C12N15/70; C12N15/73
Claims Analysis
Total Claims: 34
Claim 1
Independent Claim
86 words
A method of producing a gene product which comprises (i) culturing a temperature sensitive bacteria, which bacteria: (a) expresses the gene product intracellularly, (b) is a lysogen defective in excision and replication functions and (c) contains within the prophage DNA sequence a temperature sensitive phage repressor gene and functional phage lysozyme-encoding genes, under permissive conditions such that the gene product is expressed intracellularly and the lysozyme-encoding genes are repressed and then (ii) raising the temperature to produce restrictive conditions such that the lysozyme-encoding genes are expressed.
Claim 2
Independent Claim
86 words
A method of producing a gene product which comprises (i) culturing a temperature sensitive bacteria, which bacteria: (a) expresses the gene product intracellularly, (b) is a lysogen defective in excision and replication functions and (c) contains within the prophage DNA sequence a temperature sensitive phage repressor gene and functional phage lysozyme-encoding genes, under permissive conditions such that the gene product is expressed intracellularly and the lysozyme-encoding genes are repressed and then (ii) raising the temperature to produce restrictive conditions such that the lysozyme-encoding genes are expressed.
Claim 3
Dependent Claim
18 words
The method of claim 1 wherein the temperature-sensitive phage repressor gene is a temperature sensitive lambda cI gene.
Claim 4
Dependent Claim
13 words
The method of claim 2 wherein the cI gene is the cI857 mutant.
Claim 5
Dependent Claim
14 words
The method of claim 2 wherein the bacteria is an E. coli lambda lysogen.
Claim 6
Dependent Claim
13 words
The method of claim 4 wherein the bacteria is E. coli strain UC5822.
Claim 7
Dependent Claim
41 words
The method of claim 4 wherein the lambda prophage DNA sequence is deleted in the genes lying between map positions 58 and 71 and is mutated in the O and P genes resulting in loss of O and P gene functions.
Claim 8
Dependent Claim
35 words
The method of claim 6 wherein the lambda prophage DNA sequence is flanked by a selectable marker which is a gene coding for a selectable trait and the O and P mutations are point mutations.
Claim 9
Dependent Claim
36 words
The method of claim 7 wherein the lambda prophage DNA sequence is flanked on the upstream end by the tn10 transposable tetracycline resistance element and the O and P genes are the O29 and P3 genes.
Claim 10
Dependent Claim
14 words
The method of claim 7 wherein the bacteria is an E. coli MG strain.
Claim 11
Dependent Claim
11 words
The method of claim 7 wherein the bacteria is strain MG3.
Claim 12
Dependent Claim
23 words
The method of claim 6 wherein the lambda Prophage DNA sequence is deleted in the genes lying between map positions 3 and 71.
Claim 13
Dependent Claim
35 words
The method of claim 11 wherein the lambda prophage DNA sequence is flanked by a selectable marker which is a gene coding for a selectable trait and the O and P mutations are point mutations.
Claim 14
Dependent Claim
36 words
The method of claim 12 wherein the lambda prophage DNA sequence is flanked on the upstream end by the tn10 transposable tetracycline resistance element and the O and P genes are the O29 and P3 genes.
Claim 15
Dependent Claim
14 words
The method of claim 12 wherein the bacteria is an E. coli MG4 strain.
Claim 16
Dependent Claim
125 words
A DNA fragment comprising (i) a selectable marker which is a gene coding for a selectable trait; (ii) a lambda prophage DNA sequence having a temperature sensitive cI repressor gene and functional lysozyme encoding genes such that the lysozyme-encoding genes are repressed under permissive conditions and expressed under restrictive conditions, wherein the prophage DNA sequence includes functional N, Q, S and R genes, is substantially deleted in the genes lying between map positions 58 and 71 and has mutations in the O and P genes resulting in loss of O and P gene functions; and, (iii) flanking DNA sequences homologous to a contiguous sequence in the chromosome of a host bacterial cell to permit recombination between the fragment and the host cell chromosome to occur.
Claim 17
Dependent Claim
125 words
A DNA fragment comprising (i) a selectable marker which is a gene coding for a selectable trait; (ii) a lambda prophage DNA sequence having a temperature sensitive cI repressor gene and functional lysozyme encoding genes such that the lysozyme-encoding genes are repressed under permissive conditions and expressed under restrictive conditions, wherein the prophage DNA sequence includes functional N, Q, S and R genes, is substantially deleted in the genes lying between map positions 58 and 71 and has mutations in the O and P genes resulting in loss of O and P gene functions; and, (iii) flanking DNA sequences homologous to a contiguous sequence in the chromosome of a host bacterial cell to permit recombination between the fragment and the host cell chromosome to occur.
Claim 18
Dependent Claim
36 words
The DNA fragment of claim 15 wherein the cI gene is the cI857 gene, the O and P mutants are the O29 and P3 mutants and the selectable marker is the tn10 transposable tetracycline resistance element.
Claim 19
Dependent Claim
24 words
The DNA fragment of claim 15 wherein the prophage DNA sequence is substantially deleted in the genes lying between map positions 3 and 71.
Claim 20
Dependent Claim
36 words
The DNA fragment of claim 17 wherein the cI gene is the cI857 gene, the O and P mutants are the O29 and P3 mutants and the selectable marker is the tn10 transposable tetracycline resistance element.
Claim 21
Dependent Claim
9 words
A bacteria comprising the DNA fragment of claim 15.
Claim 22
Dependent Claim
9 words
A bacteria comprising the DNA fragment of claim 17.
Claim 23
Dependent Claim
10 words
The bacteria of claim 19 which is an E. coli.
Claim 24
Dependent Claim
10 words
The bacteria of claim 20 which is an E. coli.
Claim 25
Dependent Claim
10 words
The bacteria of claim 21 which is an MG strain.
Claim 26
Dependent Claim
9 words
The bacteria of claim 21 which is strain MG3.
Claim 27
Dependent Claim
9 words
The bacteria of claim 22 which is strain MG4.
Claim 28
Independent Claim
139 words
A method of making a temperature sensitive bacteria which comprises transforming a bacteria with a DNA fragment comprising (i) a selectable marker which is a gene coding for a selectable trait; (ii) a lambda prophage DNA sequence having a temperature sensitive cI repressor gene and functional lysozyme encoding genes such that the lysozyme-encoding genes are repressed under permissive conditions and expressed under restrictive conditions, wherein the prophage DNA sequence includes functional N, Q, S and R genes, is substantially deleted in the genes lying between map positions 58 and 71 and has mutations in the O and P genes resulting in loss of O and P gene functions; and, (iii) flanking DNA sequences homologous to a contiguous sequence in the chromosome of a host bacterial cell to permit recombination between the fragment and the host cell chromosome to occur.
Claim 29
Independent Claim
139 words
A method of making a temperature sensitive bacteria which comprises transforming a bacteria with a DNA fragment comprising (i) a selectable marker which is a gene coding for a selectable trait; (ii) a lambda prophage DNA sequence having a temperature sensitive cI repressor gene and functional lysozyme encoding genes such that the lysozyme-encoding genes are repressed under permissive conditions and expressed under restrictive conditions, wherein the prophage DNA sequence includes functional N, Q, S and R genes, is substantially deleted in the genes lying between map positions 58 and 71 and has mutations in the O and P genes resulting in loss of O and P gene functions; and, (iii) flanking DNA sequences homologous to a contiguous sequence in the chromosome of a host bacterial cell to permit recombination between the fragment and the host cell chromosome to occur.
Claim 30
Dependent Claim
35 words
The method of claim 26 wherein the cI gene is the cI857 gene, the O and P mutants are the O29 and P3 mutants and the selectable marker is the tn10 transposable tetracycline resistance element.
Claim 31
Dependent Claim
23 words
The method of claim 26 wherein the prophage DNA sequence is substantially deleted in the genes lying between map positions 3 and 71.
Claim 32
Dependent Claim
35 words
The method of claim 28 wherein the cI gene is the cI857 gene, the O and P mutants are the O29 and P3 mutants and the selectable marker is the tn10 transposable tetracycline resistance element.
Claim 33
Dependent Claim
12 words
The method of claim 26 wherein the bacteria is an E. coli.
Claim 34
Dependent Claim
12 words
The method of claim 28 wherein the bacteria is an E. coli.
References Cited (Backward Citations)
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| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
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| 3.5 Years (Year 4) | 1990-01-19 | 1990-07-21 | 1991-01-20 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1994-01-19 | 1994-07-21 | 1995-01-20 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1998-01-19 | 1998-07-21 | 1999-01-20 | $7700.00 | Window Closed / Lapsed |