Isolation and Characterization of Murine Transcriptional Control Elements Using a "Shotgun" Method

Publisher Website
Google Scholar
Abstract
In an attempt to identify novel promoter/enhancer elements in cellular genomes, an experimental "shotgun" system was designed and tested. Fragments of mouse cellular DNA were cloned into various plasmid vectors whose common reporter gene facilitates the identification of regulatory sequences. Of 120 clones analyzed, we identified one whose insert markedly increases transcriptional activity of the constructions. This genomic fragment contains a promoter element and exhibits enhancer-like properties in fibroblast cell lines of different species. Genomic analysis shows the fragment is a member of a middle repetitive sequence family, containing a 170-bp sequence with extreme purine/pyrimidine asymmetry. This stretch of cellular DNA is very sensitive to S1 nuclease digestion when present in superhelical plasmids. Isolation and characterization of sequences homologous to this fragment suggest a relationship between transcriptional activity and S1 sensitivity.