Patent EP0338841A1 successfully refreshed from Google Patents.
EP0338841A1
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EP
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Recombinant DNA methods, vectors and host cells
Assignee: Lonza Group AG
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Filing: 1989-04-18
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Granted: —
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Expires: 2009-04-18
Abstract
Abstract The present invention relates to a method for transforming a lymphoid cell line to glutamine independence which comprises: transforming the lymphoid cell line with a vector containing an active glutamine synthetase (GS) gene; growing the transformed cell line on a medium containing glutamine; and continuing the growth of the transformed cell line on a medium in which the glutamine is progressively depleted or on a medium lacking glutamine, and to vectors and host cells for use in the method.
Inventors
- 👤 Christopher Robert Bebbington
- 👤 Geoffrey Thomas Yarranton
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-03 20:31
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/93; C12N15/00; C12N15/09; C12N15/63; C12N15/67; C12N15/79; C12N15/85; C12N9/14; C12N9/48; C12N9/50; C12N9/64; C12N9/6421; C12N9/6424; C12N9/6456; C12N9/6459; C12Y; C12Y304/00; C12Y304/21; C12Y304/21069; C12N2840/00; C12N2840/20
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
63 words
A method for transforming a lymphoid cell line to glutamine independence which comprises: transforming the lymphoid cell line with a vector containing an active glutamine synthetase (GS) gene; growing the transformed cell line on a medium containing glutamine; and continuing the growth of the transformed cell line on a medium in which the glutamine is progressively depleted or on a medium lacking glutamine.
Claim 2
Independent Claim
63 words
A method for transforming a lymphoid cell line to glutamine independence which comprises: transforming the lymphoid cell line with a vector containing an active glutamine synthetase (GS) gene; growing the transformed cell line on a medium containing glutamine; and continuing the growth of the transformed cell line on a medium in which the glutamine is progressively depleted or on a medium lacking glutamine.
Claim 3
Dependent Claim
15 words
The method of claim 1, wherein the lymphoid cell line is a myeloma cell line.
Claim 4
Dependent Claim
16 words
The method of claim 1 or claim 2, wherein the glutamine-depleted or glutamine-free medium contains asparagine.
Claim 5
Independent Claim
50 words
The method of any one of claims 1 to 3, wherein the lymphoid cell line is transformed with a vector containing both an active GS gene and a gene encoding another selectable marker, such as a gpt gene, or cotransformed with separate vectors encoding GS and the selectable marker respectively.
Claim 6
Independent Claim
29 words
The method of any one of claims 1 to 4, wherein the glutamine in the medium is progressively depleted by dilution with a medium containing aspargine but lacking glutamine.
Claim 7
Independent Claim
35 words
The method of any one of claims 1 to 5, wherein the vector used to transform the lymphoid cell line also contains an active gene coding for a protein heterologous to the lymphoid cell line.
Claim 8
Independent Claim
30 words
The method of any one of claims 1 to 6, wherein the lymphoid cell line is co-transformed with a separate vector containing the active gene coding for the heterologous protein.
Claim 9
Dependent Claim
16 words
The method of claim 6 or claim 7, wherein the heterologous protein is an Ig-type molecule.
Claim 10
Independent Claim
61 words
The method of any one of claims 6 to 8, wherein the GS gene comprises a relatively weak promoter and the gene (or genes) encoding the heterologous protein comprises a relatively strong promoter so that in the transformed cell lines, protein synthesis is directed preferentially to the production of the heterologous protein or peptide rather than to the production of GS.
Claim 11
Dependent Claim
72 words
A vector for transforming a lymphoid cell line to glutamine independence and to enable it to produce a heterologous protein, the vector comprising a GS gene and a gene encoding the heterologous protein, wherein the vector is arranged such that expression of the GS gene is not hindered by transcriptional interference from the promoter/enhancer transcribing the coding sequence for the heterologous protein to such an extent that glutamine-independent colonies cannot be produced.
Claim 12
Dependent Claim
72 words
A vector for transforming a lymphoid cell line to glutamine independence and to enable it to produce a heterologous protein, the vector comprising a GS gene and a gene encoding the heterologous protein, wherein the vector is arranged such that expression of the GS gene is not hindered by transcriptional interference from the promoter/enhancer transcribing the coding sequence for the heterologous protein to such an extent that glutamine-independent colonies cannot be produced.
Claim 13
Dependent Claim
52 words
The vector of claim 10, wherein the GS gene contains a relatively weak promoter, the gene encoding the heterologous protein contains a relatively strong promoter, and the promoter of the GS gene is located upstream of or directs expression in the opposite direction to that of the gene encoding the heterologous protein.
Claim 14
Dependent Claim
23 words
The vector of claim 11, wherein the combination for the weak and strong promoters is the SV40 early region and the hCMV-MIE promoters.
Claim 15
Dependent Claim
61 words
The vector of claim 10 or claim 11, wherein the vector comprises a GS gene having a weak promoter having downstream therefrom a heavy chain like gene having a strong promoter, there being on the vector a light chain like gene having a strong promoter oriented in the opposite direction to the promoters of the GS and heavy chain like genes.
Claim 16
Dependent Claim
60 words
The vector of claim 10 or claim 11, wherein the GS gene has a weak promoter, the vector contains a light chain like gene and a heavy chain like gene, the heavy and light chain like genes have strong promoters, the three genes are transcribed in the same direction and the GS gene is upstream of the other two genes.
References Cited (Backward Citations)
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