Development of apoptosis-resistant dihydrofolate reductase-deficient Chinese hamster ovary cell line

2003 ◽  
Vol 82 (7) ◽  
pp. 872-876 ◽  
Author(s):  
Suk Kyoo Lee ◽  
Gyun Min Lee
Keyword(s):  
Cell Line ◽  
Chinese Hamster Ovary Cell ◽  
Dihydrofolate Reductase ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Ovary Cell ◽  
Ovary Cell Line

scholarly journals Organization of a Chinese hamster ovary dihydrofolate reductase gene identified by phenotypic rescue.

1983 ◽  
Vol 3 (7) ◽  
pp. 1266-1273 ◽  
Author(s):  
J D Milbrandt ◽  
J C Azizkhan ◽  
K S Greisen ◽  
J L Hamlin
Keyword(s):  
Cell Line ◽  
Chinese Hamster Ovary Cell ◽  
Dihydrofolate Reductase ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Restriction Enzymes ◽  
Ovary Cell ◽  
Dhfr Gene ◽  
Ovary Cell Line ◽  
Spheroplast Fusion

We have constructed a genomic DNA library from a methotrexate-resistant Chinese hamster ovary cell line (CHOC 400) in the cosmid vector pHC79. By utilizing a murine dihydrofolate reductase (DHFR) cDNA clone, we have identified 66 DHFR+ clones among the 11,000 colonies screened by colony hybridization. To isolate a recombinant cosmid containing the entire DHFR gene, we have tested these colonies for their ability to rescue a DHFR- Chinese hamster ovary cell line, using the spheroplast fusion method of gene transfer developed by W. Schaffner (Proc. Natl. Acad. Sci. U.S.A. 77:2163-2167, 1980). One clone (cH1) was able to transform DHFR- cells to the DHFR+ phenotype and was shown in hybridization studies to contain all of the gene except a small portion of the 3' untranslated region. We have mapped cosmid cH1 and several overlapping cosmids with a variety of restriction enzymes and have determined the approximate positions of the five (and possibly six) exons within the DHFR gene. Differences between the sizes of homologous genes in hamster cells (24.5 kilobases [kb]) and in mouse cells (31.5 kb) are shown to reside primarily in the length of the 3' intron, which is 8 kb in the hamster gene and 16 kb in length in the mouse gene. Our studies confirm the utility of cosmid libraries for the isolation of large genes, as previously shown by R. de Saint Vincent et al. (Cell 27:267-277, 1981). In addition, a cosmid that contains a functional DHFR gene will be a useful vector for the co-amplification and subsequent overexpression of other cloned genes.

Organization of a Chinese hamster ovary dihydrofolate reductase gene identified by phenotypic rescue

1983 ◽  
Vol 3 (7) ◽  
pp. 1266-1273
Author(s):  
J D Milbrandt ◽  
J C Azizkhan ◽  
K S Greisen ◽  
J L Hamlin
Keyword(s):  
Cell Line ◽  
Chinese Hamster Ovary Cell ◽  
Dihydrofolate Reductase ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Restriction Enzymes ◽  
Ovary Cell ◽  
Dhfr Gene ◽  
Ovary Cell Line ◽  
Spheroplast Fusion

We have constructed a genomic DNA library from a methotrexate-resistant Chinese hamster ovary cell line (CHOC 400) in the cosmid vector pHC79. By utilizing a murine dihydrofolate reductase (DHFR) cDNA clone, we have identified 66 DHFR+ clones among the 11,000 colonies screened by colony hybridization. To isolate a recombinant cosmid containing the entire DHFR gene, we have tested these colonies for their ability to rescue a DHFR- Chinese hamster ovary cell line, using the spheroplast fusion method of gene transfer developed by W. Schaffner (Proc. Natl. Acad. Sci. U.S.A. 77:2163-2167, 1980). One clone (cH1) was able to transform DHFR- cells to the DHFR+ phenotype and was shown in hybridization studies to contain all of the gene except a small portion of the 3' untranslated region. We have mapped cosmid cH1 and several overlapping cosmids with a variety of restriction enzymes and have determined the approximate positions of the five (and possibly six) exons within the DHFR gene. Differences between the sizes of homologous genes in hamster cells (24.5 kilobases [kb]) and in mouse cells (31.5 kb) are shown to reside primarily in the length of the 3' intron, which is 8 kb in the hamster gene and 16 kb in length in the mouse gene. Our studies confirm the utility of cosmid libraries for the isolation of large genes, as previously shown by R. de Saint Vincent et al. (Cell 27:267-277, 1981). In addition, a cosmid that contains a functional DHFR gene will be a useful vector for the co-amplification and subsequent overexpression of other cloned genes.

scholarly journals Characterization of recombinant human orexin receptor pharmacology in a Chinese hamster ovary cell-line using FLIPR

1999 ◽  
Vol 128 (1) ◽  
pp. 1-3 ◽  
Author(s):  
D Smart ◽  
J C Jerman ◽  
S J Brough ◽  
S L Rushton ◽  
P R Murdock ◽  
...  
Keyword(s):  
Cell Line ◽  
Chinese Hamster Ovary Cell ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster ◽  
Ovary Cell ◽  
Receptor Pharmacology ◽  
Ovary Cell Line
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