The eucaryotic cell-lipochromosome as a new host-vector system in gene amplification and expression.
Although procaryotes such as E. Coli are generally considered to be ideal hosts, able to amplify eucaryotic gene sequences contained within hybrid DNA plasmid molecules (3, 14), recent experimental evidence such as the decreased bacterial viability observed during the cloning of mouse mitochondrial DNA, clearly shows the limitations of this type of approach (6). In addition, major technical difficulties are encountered during the isolation and purification of the specific messenger RNA (mRNA) needed to synthetize the complementary DNA (cDNA) molecule (1, 4). Last but not least, after having screened many different bacterial clones -- provided that the foreign gene product is not toxic for its host -- it is still a question of good luck to get one clone producing adequate quantities of the biologically active protein of interest, free of contaminants. On the other hand, the low but significant frequency with which eucaryotic cells exposed to fragmented DNA molecules or metaphasic chromosomes phenotypically express a particular marker suggests that this approach might offer an alternative to the bacteria-plasmid system used in "classic" genetic engineering (10, 17, 18, 19). It is the purpose of this work to briefly discuss some of the difficulties involved in this approach and to propose solutions.
D002875:Chromosomes / Q000378:metabolism*; D003001:Cloning, Molecular; D046508:Culture Techniques / Q000379:methods; D004247:DNA; D004926:Escherichia coli / Q000235:genetics; D005784:Gene Amplification; D005818:Genetic Engineering; D010957:Plasmids; D014158:Transcription, Genetic
D004247:DNA