Patent EP0057976A2 successfully refreshed from Google Patents.
EP0057976A2
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A process for cloning the gene coding for a thermostable alpha-amylase into escherichia coli and bacillus subtilis
Assignee: Unilever Bestfoods North America
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Filing: 1982-01-12
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Granted: —
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Expires: 2002-01-12
Abstract
Abstract An improved process for producing a thermostable alpha-amylase enzyme is described. The gene coding for the alpha-amylase is incorporated into a chimeric plasmid which is produced in multiple copies by a host microorganism.
Inventors
- 👤 Jonathan R. Mielenz
- 👤 Susan Mickel
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-29 20:24
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/14; C12N9/24; C12N9/2402; C12N9/2405; C12N9/2408; C12N9/2411; C12N9/2414; C12N9/2417
Claims Analysis
Total Claims: 26
Claim 1
Independent Claim
51 words
A chimeric plasmid having a gene coding for a thermostable alpha-amylase prepared by cutting DNA from a donor and combining the resulting DNA fragments with a vector which has been similarly cut, wherein the improvement comprises using as a donor a naturally-occurring plasmid containing a gene coding for a thermostable alpha-amylase.
Claim 2
Dependent Claim
51 words
A chimeric plasmid having a gene coding for a thermostable alpha-amylase prepared by cutting DNA from a donor and combining the resulting DNA fragments with a vector which has been similarly cut, wherein the improvement comprises using as a donor a naturally-occurring plasmid containing a gene coding for a thermostable alpha-amylase.
Claim 3
Dependent Claim
18 words
The chimeric plasmid of claim 1 wherein the donor plasmid is derived from a strain of B. stearothermophilus.
Claim 4
Dependent Claim
39 words
The chimeric plasmid of claim 2 wherein the strain of B. stearothermophilus is selected from the group consisting of B. stearothermophilus, ATCC Nos. 31,195; 31,196; 31,197; 31,198; 31,199; and 31,783, variants and mutants thereof and submutants of said mutants.
Claim 5
Dependent Claim
33 words
The chimeric plasmid of claims 1, 2 or 3 wherein the vector is a plasmid selected from the group consisting of E. coli plasmids pBR322, pBR325 and pWL625; and B. subtilis plasmid pC194.
Claim 6
Dependent Claim
25 words
A microorganism selected from the group comprising E. coli and B. subtilis characterized in that it comprises at least one chimeric plasmid of claim 1.
Claim 7
Dependent Claim
65 words
A process for constructing a chimeric plasmid having a gene coding for a thermostable alpha-amylase which comprises: (a) cutting naturally-occurring plasmid DNA having a gene coding for a thermostable alpha-amylase to obtain a linear DNA sequence containing the alpha-amylase coding gene; (b) cutting a suitable vector to obtain a second linear DNA sequence; and (c) joining the linear DNA sequences of Steps (a) and (b).
Claim 8
Dependent Claim
65 words
A process for constructing a chimeric plasmid having a gene coding for a thermostable alpha-amylase which comprises: (a) cutting naturally-occurring plasmid DNA having a gene coding for a thermostable alpha-amylase to obtain a linear DNA sequence containing the alpha-amylase coding gene; (b) cutting a suitable vector to obtain a second linear DNA sequence; and (c) joining the linear DNA sequences of Steps (a) and (b).
Claim 9
Dependent Claim
29 words
The process of claim 6 wherein the vector is a plasmid selected from the group consisting of E. coli plasmids pBR322, pBR325 and pWL625; and B. subtilis plasmid pC194.
Claim 10
Dependent Claim
19 words
The process of claim 6 characterized in that the naturally-occurring plasmid is derived from a strain of B. stearothermophilus.
Claim 11
Dependent Claim
38 words
The process of claim 8 wherein the strain of B. stearothermophilus is selected from the group consisting of B. stearothermophilus, ATCC Nos. 31,195; 31,196; 31,-197; 31,198; 31,199; and 31,783, variants and mutants thereof and submutants of said mutants.
Claim 12
Dependent Claim
32 words
The process of claim 6 characterized in that the cutting is accomplished by means of the restriction endonuclease, Hind III, and the joining is accomplished by means of T 4 DNA ligase.
Claim 13
Dependent Claim
49 words
A process for preparing a thermostable alpha-amylase which comprises the steps of: (a) introducing at least one chimeric plasmid of claim 1 into a host microorganism; (b) culturing the microorganism containing the chimeric plasmid in a suitable medium; and (c) isolating the alpha amylase produced by the cultured microorganism.
Claim 14
Dependent Claim
65 words
A chimeric plasmid having a gene coding for a thermostable alpha-amylase prepared by cutting DNA from a donor and combining the resulting DNA fragments with a vector which has been similarly cut, wherein the improvement comprises using as a donor a different chimeric plasmid, containing a DNA fragment with a gene coding for a thermostable alpha-amylase, said DNA fragment being obtained from a naturally-occurring plasmid.
Claim 15
Dependent Claim
65 words
A chimeric plasmid having a gene coding for a thermostable alpha-amylase prepared by cutting DNA from a donor and combining the resulting DNA fragments with a vector which has been similarly cut, wherein the improvement comprises using as a donor a different chimeric plasmid, containing a DNA fragment with a gene coding for a thermostable alpha-amylase, said DNA fragment being obtained from a naturally-occurring plasmid.
Claim 16
Dependent Claim
18 words
The chimeric plasmid of claim 12 wherein the naturally-occurring plasmid is derived from a strain of B. stearothermophilus.
Claim 17
Dependent Claim
39 words
The chimeric plasmid of claim 13 wherein the strain of B. stearothermophilus is selected from the group consisting of B. stearcthermophilus, ATCC Nos. 31,195; 31,196; 31,197; 31,198; 31,199; and 31,783, variants and mutants thereof and submutants- of said mutants.
Claim 18
Dependent Claim
33 words
The chimeric plasmid of claims 12, 13 or 14 wherein the vector is a plasmd selected from the group consisting of E. coli plasmids pBR322, p3R325 and pWL625; and B. subtilis plasmid pC194.
Claim 19
Dependent Claim
25 words
A microorganism selected from the group comprising E. coli and B. subtilis characterized in that it comprises at least one chimeric plasmid of claim 12.
Claim 20
Dependent Claim
79 words
A process for constructing a chimeric plasmid having a gene coding for a thermostable alpha-amylase which comprises: (a) cutting a different chimeric plasmid, containing a DNA fragment with a gene coding for a thermostable alpha-amylase, said DNA fragment being obtained from a naturally-occurring plasmid, to obtain a linear DNA sequence containing the alpha-amylase coding gene; (b) cutting a suitable vector to obtain a second linear DNA sequence; and (c) joining the linear DNA sequences of Steps (a) and (b).
Claim 21
Dependent Claim
79 words
A process for constructing a chimeric plasmid having a gene coding for a thermostable alpha-amylase which comprises: (a) cutting a different chimeric plasmid, containing a DNA fragment with a gene coding for a thermostable alpha-amylase, said DNA fragment being obtained from a naturally-occurring plasmid, to obtain a linear DNA sequence containing the alpha-amylase coding gene; (b) cutting a suitable vector to obtain a second linear DNA sequence; and (c) joining the linear DNA sequences of Steps (a) and (b).
Claim 22
Dependent Claim
29 words
The process of claim 17 wherein the vector is a plasmid selected from the group consisting of E. coli plasmids pBR322, pBR325 and pWL625; and B. subtilis plasmid pC194.
Claim 23
Dependent Claim
19 words
The process of claim 17 characterized in that the naturally-occurring plasmid is derived from a strain of B. stearothermophilus.
Claim 24
Dependent Claim
38 words
The process of claim 19 wherein the strain of B. stearothermophilus is selected from the group consisting of 3. stearothermophilus, ATCC Nos. 31,195; 31,196; 31,197; 31,198; 31,199; and 31,783, variants and mutants thereof and submutants of said mutants.
Claim 25
Dependent Claim
32 words
The process of claim 17 characterized in that the cutting is accomplished by means of the restriction endonuclease, Hind III, and the joining is accomplished by means of T 4 DNA ligase.
Claim 26
Dependent Claim
48 words
A process for preparing a thermostable alpha-amylase which comprises the steps of: (a) introducing at least one chimeric plasmid of claim 12 into a host microorganism; (b) culturing the microorganism containing the chimeric plasmid in a suitable medium; and (c) isolating the alpha-amylase produced by the cultured microorganism.
References Cited (Backward Citations)
No citation records indexed.
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