Cotransfection and cointegration of pAdNLK and pAdD26SVp(A)3 enables the SV40 enhancer element of pAdNLK to activate the mouse dihydrofolate reductase transcription unit of pAdD26SVp(A)3. This cell line was produced by cotransfection with neuroleukin and dihydrofolate reductase transcription units. Production of neuroleukin was increased by selection with methotrexate. Recently, it has been shown that the polypeptide neuroleukin is in fact glucophosphoisomerase. pAdNLK contains the SV40 enhancer, the mouse neuroleukin cDNA, the mouse dihydrofolate reductase cDNA, the SV40 polyadenylation sequence, and pBR322 sequences minus the inhibitory sequences from 1424 to 2490. pAdD26SVp(A)3 contains the adenovirus major late promoter (AdMLP), the mouse dihydrofolate reductase cDNA, the SV40 polyadenylation sequence, pBR322 sequences minus the inhibitory sequences from 1424 to 2490, and the SV40 origin of replication. The pAdNLK insert contains the complete nucleotide sequence and 5' and 3' untranslated sequences. The cell line CHO-IC6 also contains pAdD26SVp(A)3. Selection of the parental line for growth in methotrexate (MTX) coamplified MTX resistance and neuroleukin secretion. CHO-IC6 secretes 4.4 ug neuroleukin per 10(7) cells per 24 hr. pAdD26SVp(A)3 was constructed by cloning the mouse dihydrofolate reductase cDNA from the plasmid pAdD26-1 into the vector, pSVOd. A 900 bp BclI/PstI fragment of SV40 containing the early polyadenylation site was inserted. |
