Patent GB2118947A successfully refreshed from Google Patents.
GB2118947A
Expired / Lapsed
GB
⏱️ 0.0 years remaining
Cloning vectors
Assignee: Eli Lilly and Co
•
Filing: 1983-04-14
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Granted: —
•
Expires: 2003-04-14
Abstract
Abstract There is described a recombinant DNA cloning vector useful in Streptomyces and E. coli, comprising a) a functional origin of replication- containing restriction fragment of plasmid SCP2 or SCP2*, b) a restriction fragment comprising an E. coli origin of replication, c) one or more DNA segments that confer resistance to at least one antibiotic when transformed into a cell of E. coli, said cell being sensitive to the antibiotic for which resistance is conferred, and d) one or more DNA segments that independently confer either or both of the Streptomyces tra function or resistance to at least one antibiotic when transformed into a cell of Streptomyces, said cell being sensitive to the antibiotic for which resistance is conferred. Transformants and a method for detecting transformants of the vector are also disclosed.
Inventors
- 👤 Charles Lee Hershberger
- 👤 Jeffrey Lynn Larson
Bibliographic Details
Application Number
—
Jurisdiction
GB
Data Source
google_patents
Last Synchronized
2026-11-08 04:30
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/63; C12N15/74; C12N15/76
Claims Analysis
Total Claims: 51
Claim 1
Independent Claim
108 words
A recombinant DNA cloning vector comprising:
a) a functional origin of replication-containing restriction fragment of plasmid SCP2 or SCP2*,
b) a restriction fragment comprising an E. coli origin of replication,
c) one or more DNA segments that confer resistance to at least one antibiotic when transformed into a cell of E. coli, said cell being sensitive to the antibiotic for which resistance is conferred, and
d) one or more DNA segments that independently confer either or both of the Streptomyces tra function or resistance to at least one antibiotic when transformed into a cell of Streptomyces, said cell being sensitive to the antibiotic for which resistance is conferred.
Claim 2
Dependent Claim
108 words
A recombinant DNA cloning vector comprising:
a) a functional origin of replication-containing restriction fragment of plasmid SCP2 or SCP2*,
b) a restriction fragment comprising an E. coli origin of replication,
c) one or more DNA segments that confer resistance to at least one antibiotic when transformed into a cell of E. coli, said cell being sensitive to the antibiotic for which resistance is conferred, and
d) one or more DNA segments that independently confer either or both of the Streptomyces tra function or resistance to at least one antibiotic when transformed into a cell of Streptomyces, said cell being sensitive to the antibiotic for which resistance is conferred.
Claim 3
Dependent Claim
9 words
The vector of Claim 1 which is a plasmid.
Claim 4
Dependent Claim
35 words
The vector of Claim 1 or 2, wherein the restriction fragment of plasmid SCP2 or SCP2* is the #5.4 kb EcoRI-Sa# fragment, #6.0 kb Sa# fragment, #19 kb EcoRI-HindIII fragment, or #31 kb EcoRI fragment.
Claim 5
Dependent Claim
38 words
The vector of Claim 1, 2 or 3, wherein the E. coli origin of replication is the pBR322 origin of
replication, pBR324 origin of replication, pBR325 origin of replication, pBR327 origin of replication, or
pBR328 origin of replication.
Claim 6
Dependent Claim
33 words
The vector of any of Claims 1 to 4, wherein the one or more DNA segments that confer
resistance in E. coli are DNA segments that confer resistance to ampicillin, chloramphenicol or tetracycline.
Claim 7
Dependent Claim
31 words
The vector of any of Claims 1 to 5, wherein the one or more DNA segments that confer
resistance in Streptomyces are DNA segments that confer resistance to neomycin or thiostrepton.
Claim 8
Dependent Claim
51 words
The vector of Claim 6 which is plasmid pJL1 80, pJL181, pJL1 25, pJL1 90, pJL1 92, pJL1 95, pJL199, pJL122, pJL123, pJL124, pJL126, pJL176, pJL1200, pJL1201, pJL1020, pJL1203, pJL1204, pJL1205, pJL1206, pJL1706, pJL1800, pJL1801, pJL1900, pJL1902, pJL1905, pJL193, pJL1 96, pJL1 97, pJL1 98, pHJL212, pHJL21 3, or pJL1 907.
Claim 9
Dependent Claim
69 words
The vector of any of Claims 1 to 7, wherein the DNA segment that confers resistance to an antibiotic is the #7.7 kb EcooRI-HindIII restriction fragment of plasmid plasmid pJL192, the #7.7 kb EcoRI
HindIII restriction fragment of plasmid pLR4, the #7.5 kb EcoRI-partial Sa# restriction fragment of plasmid pLR4, the #1.35 kb BamHI restriction fragment of plasmid pLR2, or the #1 kb Bc# restriction fragment of plasmid pJL193.
Claim 10
Dependent Claim
30 words
The vector of Claim 8 which is a plasmid pJL1 90 or pJL195 high resistance mutant that in
Streptomyces confers resistance to neomycin at levels of at least 10 g/ml.
Claim 11
Dependent Claim
3 words
Plasmid pJL1 14.
Claim 12
Dependent Claim
3 words
Plasmid pJL1 14.
Claim 13
Dependent Claim
3 words
Plasmid pJL1 20.
Claim 14
Dependent Claim
3 words
Plasmid pJL1 20.
Claim 15
Dependent Claim
3 words
Plasmid pJL1 21.
Claim 16
Dependent Claim
3 words
Plasmid pJL1 21.
Claim 17
Dependent Claim
2 words
Plasmid pJL192.
Claim 18
Dependent Claim
2 words
Plasmid pJL192.
Claim 19
Dependent Claim
2 words
Plasmid pJL193.
Claim 20
Dependent Claim
2 words
Plasmid pJL193.
Claim 21
Dependent Claim
3 words
Plasmid pJL1 97.
Claim 22
Dependent Claim
3 words
Plasmid pJL1 97.
Claim 23
Dependent Claim
3 words
Plasmid pJL1 98.
Claim 24
Dependent Claim
3 words
Plasmid pJL1 98.
Claim 25
Dependent Claim
2 words
Plasmid pHJL212.
Claim 26
Dependent Claim
2 words
Plasmid pHJL212.
Claim 27
Dependent Claim
2 words
Plasmid pHJL213.
Claim 28
Dependent Claim
2 words
Plasmid pHJL213.
Claim 29
Dependent Claim
17 words
A transformed host cell comprising the recombinant DNA cloning vector of any of Claims 1 to 18.
Claim 30
Dependent Claim
17 words
The host cell of Claim 19 which is Streptomyces, preferably of species lividans, griseofuscus, fradiae, or ambofaciens.
Claim 31
Dependent Claim
11 words
The host cell of Claim 19 which is E. coli K12.
Claim 32
Dependent Claim
21 words
A restriction fragment comprising the plasmid SCP2 or SCP2* #5.4 kb EcoRI-Sa# or #6.0 kb Sa# restriction fragment of Claim 3.
Claim 33
Dependent Claim
16 words
A restriction fragment comprising the plasmid pJL1 92 #7.7 kb EcoRI-Hindlil restriction fragment of Claim 8.
Claim 34
Dependent Claim
120 words
A process for preparing a recombinant DNA cloning vector which comprises ligating a functional origin of replication-containing restriction fragment of plasmid SCP2 or SCP2* and one or more DNA sequences comprising:
a) a restriction fragment comprising an E. coli origin of replication,
b) one or more DNA segments that confer resistance to at least one antibiotic when transformed into a cell of E. coll, said cell being sensitive to the antibiotic for which resistance is conferred, and
c) one or more DNA segments that independently confer either or both of the Streptomyces tra function or resistance to at least one antibiotic when transformed into a cell of Streptomyces, said cell being sensitive to the antibiotic for which resistance is conferred.
Claim 35
Dependent Claim
120 words
A process for preparing a recombinant DNA cloning vector which comprises ligating a functional origin of replication-containing restriction fragment of plasmid SCP2 or SCP2* and one or more DNA sequences comprising:
a) a restriction fragment comprising an E. coli origin of replication,
b) one or more DNA segments that confer resistance to at least one antibiotic when transformed into a cell of E. coll, said cell being sensitive to the antibiotic for which resistance is conferred, and
c) one or more DNA segments that independently confer either or both of the Streptomyces tra function or resistance to at least one antibiotic when transformed into a cell of Streptomyces, said cell being sensitive to the antibiotic for which resistance is conferred.
Claim 36
Dependent Claim
111 words
The process of Claim 24, wherein the restriction fragment of plasmid SCP2 or SCP2* is the 5.4 kb EcoRI-Sail fragment, ""6.0 kb Sail fragment, ""19 kb EcoRI-Hindill fragment, or ""31 kb EcoRI fragment and wherein the E. collorigin of replication is the pBR322 origin of replication, pBR324 origin of replication, pBR325 origin of replication, pBR327 origin of replication, or pBR328 origin of replication, and wherein the one or more DNA segments that confer resistance in E. coli are DNA segments that confer resistance to ampicillin, chloramphenicol or tetracycline and wherein the one or more DNA segments that confer resistance in Streptomyces are DNA segments that confer resistance to neomycin or thiostrepton.
Claim 37
Independent Claim
76 words
A method for detecting transformants comprising:
1) mixing Streptomyces cells, under transforming conditions, with a recombinant DNA cloning vector, said vector comprising
a) an origin of replication and P gene-containing restriction fragment of plasmid SCP2 or SCP2*,
and
b) a non-lethal DNA sequence cloned into the EcoRI restriction site of said P gene, and
2) growing said Streptomyces cells on a lawn of an indicator Streptomyces strain and selecting colonies that show the M pock phenotype.
Claim 38
Independent Claim
76 words
A method for detecting transformants comprising:
1) mixing Streptomyces cells, under transforming conditions, with a recombinant DNA cloning vector, said vector comprising
a) an origin of replication and P gene-containing restriction fragment of plasmid SCP2 or SCP2*,
and
b) a non-lethal DNA sequence cloned into the EcoRI restriction site of said P gene, and
2) growing said Streptomyces cells on a lawn of an indicator Streptomyces strain and selecting colonies that show the M pock phenotype.
Claim 39
Dependent Claim
22 words
The method of Claim 26, wherein the restriction fragment comprising the origin of replication and the P gene is of plasmid SCP2*.
Claim 40
Dependent Claim
23 words
The method of Claim 26 or 27, wherein the Streptomyces cells and indicator strain are preferably of species lividans, griseofuscus, fradiae or ambofaciens.
Claim 41
Dependent Claim
37 words
The method of Claim 26, 27 or 28, wherein the recombinant DNA cloning vector is plasmid pJL1 20, pJL1 21, pJL1 25, pJL1 90, pJL1 92, pJL1 95, pJL1 97, pJL1 98, pHJL21 2, or pHJL21 3.
Claim 42
Dependent Claim
24 words
A recombinant DNA cloning vector substantially as hereinbefore described with particular reference to Examples 8, 10, 12, 15, 26, 27 and 30 to 56.
Claim 43
Dependent Claim
24 words
A recombinant DNA cloning vector substantially as hereinbefore described with particular reference to Examples 8, 10, 12, 15, 26, 27 and 30 to 56.
Claim 44
Dependent Claim
15 words
A restriction fragment substantially as hereinbefore described with particular reference to
Examples 8a and 12b.
Claim 45
Dependent Claim
15 words
A restriction fragment substantially as hereinbefore described with particular reference to
Examples 8a and 12b.
Claim 46
Dependent Claim
24 words
A transformed host cell substantially as hereinbefore described with particular reference to
Examples 9, 11, 13, 16, and 19 to 25, and Table 2.
Claim 47
Dependent Claim
24 words
A transformed host cell substantially as hereinbefore described with particular reference to
Examples 9, 11, 13, 16, and 19 to 25, and Table 2.
Claim 48
Dependent Claim
28 words
A process for preparing a recombinant DNA cloning vector substantially as hereinbefore described with particular reference to Examples 8, 10, 12, 15, 26, 27 and 30 to 56.
Claim 49
Dependent Claim
28 words
A process for preparing a recombinant DNA cloning vector substantially as hereinbefore described with particular reference to Examples 8, 10, 12, 15, 26, 27 and 30 to 56.
Claim 50
Independent Claim
15 words
A method for detecting transformants substantially as hereinbefore described with particular reference to Example 19C.
Claim 51
Independent Claim
15 words
A method for detecting transformants substantially as hereinbefore described with particular reference to Example 19C.
References Cited (Backward Citations)
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