Cloning of DNA complementary to rat liver fatty acid synthetase mRNA

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Abstract
Clones, containing DNA complementary (cDNA) to rat liver fatty acid synthetase mRNA, were constructed and identified, cDNA of these clones was then used as a probe to quantify mRNA. The cDNA was synthesized to partially purified rat liver fatty acid synthetase mRNA. Double-stranded cDNA was then prepared and inserted into the PstI site of pBR322 using oligo(dG) .cntdot. oligo(dC) tailing. Initial selection of the clones was by differential colony hybridization employing [32P]cDNA synthesized from poly(A)-rich mRNA, enriched and non-enriched in fatty acid synthetase mRNA, as probes. Plasmids, containing specific sequences complementary to the fatty acid synthetase mRNA, were identified by hybrid-arrest translation. Cloned cDNA inserts ranged from 300-1400 base pairs. Cloned cDNA was employed to probe for mRNA in hybridizations via the dot-blot method. An increase in fatty acid synthetase mRNA during dietary induction was demonstrated, which suggests that regulation may involve changes in transcription or changes in post-transcriptional processing of the mRNA.