Isolation of growth suppressors from a cDNA expression library.
We describe an experimental procedure for the isolation of growth inhibitory sequences from a complex cDNA library. This approach first takes advantage of the SETGAP technique (selectable expression of transient growth arrest phenotype) to enrich for growth inhibitory sequences, followed by a screening procedure to identify individual cDNAs that inhibit cell proliferation. Here we provide a detailed description of the experimental protocol and report the characterization of two cDNA sequences isolated in our initial screen of a mouse cDNA library. One of these cDNAs encodes the mouse ubiquitin-conjugation enzyme UbcM2. The other encodes a truncated form of a novel WD40 repeat protein, named Bopl, which is conserved from yeast to human. Together, these results demonstrate a new approach for the isolation of growth suppressors from cDNA libraries, and identify a previously unknown gene likely to be involved in growth control.
Pestov
D G
DG
Department of Molecular Genetics, University of Illinois at Chicago College of Medicine, 60607-7170, USA.
Grzeszkiewicz
T M
TM
Lau
L F
LF
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D018076:DNA, Complementary; D005656:Fungal Proteins / Q000235:genetics*; D016193:G1 Phase; D015870:Gene Expression; D015723:Gene Library; D006131:Growth Inhibitors / Q000235:genetics*; D006801:Humans; D008025:Ligases / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D044763:Ubiquitin-Conjugating Enzymes
D018076:DNA, Complementary; C116590:FLC1 protein, S cerevisiae; D005656:Fungal Proteins; D006131:Growth Inhibitors; D029701:Saccharomyces cerevisiae Proteins; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases