SUPPRESSION OF ANCHORAGE-INDEPENDENT GROWTH AFTER GENE TRANSFECTION

A novel procedure for isolating anchorage-dependent cells has been dev eloped- It involves negative selection of cells growing in suspension followed by clonal replica screening for anchorage-dependent growth. C ells which have regained anchorage-dependent growth have been isolated from a library of the Chinese hamster ovary cell line, CHO-KI, transf ected with pSV2neo and human genomic DNA. One anchorage-dependent clon e, 1042AC, has been studied in detail. Anchorage-dependent growth of 1 042AC is stable when cultured as adherent monolayers, but revertants a ppear rapidly when cultured in suspension. Suppression is unlikely to be due to loss or mutation of hamster genes conferring anchorage-indep endent growth as hybrids between 1042AC and CHO-K1 have the suppressed phenotype of 1042AC. Furthermore, a population of cells obtained from the hybrid by selecting for revertants to anchorage-independent growt h showed selective loss of the transgenome derived from 1042AC. The gr owth suppression was not due to transfection of the human Krev-1 gene, which has previously been shown to restore anchorage-dependent growth , nor was there any evidence of alteration in the endogenous hamster K rev-1 gene. However, evidence for a human gene being responsible for t he suppressed phenotype has not been obtained yet.