EP0260148A2
Expired / Lapsed
EP
⏱️ 0.0 years remaining
Improved recombinant expression method, vector and transformed cells
Assignee: Genentech Inc
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Filing: 1987-09-11
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Granted: —
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Expires: 2007-09-11
Abstract
Abstract A method for continuous production of a desired heterologous protein comprising constructing an expression vector having a stabilizing sequence downstream of a promoter and upstream of the DNA encoding the desired heterologous protein, transfecting and choosing a particular eukaryotic host cell for said continuous production and culturing the transformed eukaryotic host cell under conditions favorable for continuous production of said desired heterologous protein.
Inventors
- 👤 Cornelia Maxine Gorman
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-03 08:49
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/14; C12N9/48; C12N9/50; C12N9/64; C12N9/6421; C12N9/6424; C12N9/6456; C12N9/6459; C07; C07K; C07K14/00; C07K14/435; C07K14/575; C07K14/64; C07K14/745; C07K14/755; C12N15/00; C12N15/09; C12N15/63; C12N15/67; C12N15/79; C12N15/85; C12Y; C12Y304/00; C12Y304/21; C12Y304/21069; C12N2840/00; C12N2840/44
Claims Analysis
Total Claims: 23
Claim 1
Independent Claim
118 words
A method for continuous production of a desired heterologous protein in a eukaryotic host cell comprising: a) constructing an expression vector having a sequence of double stranded DNA comprising the following elements: 1) a stabilizing sequence downstream of a promoter and upstream of a DNA encoding the amino acid sequence of the desired heterologous protein; 2) DNA encoding the amino acid sequence of the desired heterologous protein downstream of said stabilizing sequence; and, 3) DNA coding a polyadenylation sequence downstream of which is a transcription termination site; b) transfecting and then choosing a eukaryotic host cell with said expression vector; and c) culturing the transfected eukaryotic host cell under conditions favourable for continuous production of the desired protein.
Claim 2
Independent Claim
118 words
A method for continuous production of a desired heterologous protein in a eukaryotic host cell comprising: a) constructing an expression vector having a sequence of double stranded DNA comprising the following elements: 1) a stabilizing sequence downstream of a promoter and upstream of a DNA encoding the amino acid sequence of the desired heterologous protein; 2) DNA encoding the amino acid sequence of the desired heterologous protein downstream of said stabilizing sequence; and, 3) DNA coding a polyadenylation sequence downstream of which is a transcription termination site; b) transfecting and then choosing a eukaryotic host cell with said expression vector; and c) culturing the transfected eukaryotic host cell under conditions favourable for continuous production of the desired protein.
Claim 3
Dependent Claim
17 words
The method of claim 1 wherein the promoter is from the immediate early gene of human cytomegalovirus.
Claim 4
Dependent Claim
14 words
The method of claim 1 wherein the promoter is from simian virus 40 (SV40).
Claim 5
Independent Claim
27 words
The method of any one of claims 1, 2 and 3 wherein the stabilizing sequence comprises at least one but not more than two splice donor-intron-acceptor sequences.
Claim 6
Dependent Claim
24 words
The method of claim 4 wherein the splice donor sequence of the splice donor-intron-acceptor sequence is from the immediate early gene of human cytomegalovirus.
Claim 7
Dependent Claim
23 words
The method of claim 4 wherein the intron of the splice donor-intron-acceptor sequence is from the human cytomegalovirus and the immunoglobulin variable region.
Claim 8
Dependent Claim
21 words
The method of claim 4 wherein the splice acceptor sequence of the splice donor-intron-acceptor sequence corresponds to the immunoglobulin acceptor sequence.
Claim 9
Independent Claim
46 words
The method of any one of claims 1, 2 and 3 wherein the stabilizing sequence comprises an engineered DNA coding a mRNA having the same properties as a mRNA which had been subject to splicing but from which no nucleotide sequence had in fact been removed.
Claim 10
Independent Claim
24 words
The method of any one of the preceding claims wherein the DNA encoding the amino acid sequence of a heterologous protein encodes factor VIII.
Claim 11
Independent Claim
24 words
The method of any one of claims 1 to 8 wherein the DNA encoding the amino acid sequence of a heterologous protein encodes t-PA.
Claim 12
Independent Claim
24 words
The method of any one of claims 1 to 8 wherein the DNA encoding the amino acid sequence of a heterologous protein encodes prorelaxin.
Claim 13
Independent Claim
28 words
The method of any one of claims 1 to 8 wherein the DNA encoding the amino acid sequence of a heterologous protein encodes a variant of factor VIII.
Claim 14
Dependent Claim
17 words
The method of claim 12 wherein the factor VIII is resistant to cleavage by activated protein C.
Claim 15
Independent Claim
22 words
The method of any one of the preceding claims wherein the DNA coding the polyadenylation sequence is from simian virus 40 (SV40).
Claim 16
Independent Claim
18 words
The method of any one of the preceding claims wherein the host cell is mouse sertoli cell (TM4).
Claim 17
Independent Claim
18 words
The method of any one of claims 1 to 14 wherein the host cell is hepatoma cell (HTC).
Claim 18
Independent Claim
20 words
The method of any one of claims 1 to 14 wherein the host cell is human embryonic kidney cell (293).
Claim 19
Independent Claim
16 words
The method of any one of the preceding claims wherein the expression vector includes an enhancer.
Claim 20
Dependent Claim
14 words
The method of claim 18 wherein the enhancer is located upstream of the promoter.
Claim 21
Dependent Claim
18 words
The method of claim 18 or claim 19 which includes an enhancer and promoter from simian virus (SV40).
Claim 22
Dependent Claim
29 words
A vector suitable for continuous expression in a eukaryotic host cell culture of a desired heterologous protein which vector has the features defined in any of the preceding claims.
Claim 23
Dependent Claim
24 words
TM4 cells, HTC cells or 293 cells transformed with an expression vector having the features defined in any one of claims 1 to 20.
References Cited (Backward Citations)
No citation records indexed.
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