Selective amplification of DNA fragments coding for the G domain of factors IF2 and EF-Tu, two G proteins from the myxobacterium Stigmatella aurantiaca.
Two DNA fragments of the genome of the myxobacterium Stigmatella aurantiaca were selectively amplified by PCR. These fragments encode a segment of the G domain of translational initiation factor IF2 and elongation factor EF-Tu, two GTP-binding proteins. This was made possible by carefully designing the primers for this reaction to avoid the amplification of every G-domain-encoding region of the genome. The sequence of two pairs of primers was deduced from highly conserved regions, namely G1 and G3 and/or their vicinal amino acids, within each subfamily (initiation and elongation factors, respectively) of GTP-binding proteins. On the basis of the expected size, one band was selected in each experiment, cloned into a vector, and sequenced. This showed unambiguously after comparison analysis that they belong to the IF2 and EF-Tu genes, respectively. This strategy seems suitable for the amplification of a segment of any gene coding for a G protein from any origin.
Bremaud
L
L
Institut de Biologie Moléculaire et d'Ingénierie Génétique, Centre National de la Recherche Scientifique (CNRS) URA1172, Université de Poitiers, France.
Derijard
B
B
Cenatiempo
Y
Y
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D017931:DNA Primers; D019204:GTP-Binding Proteins / Q000235:genetics*; D005784:Gene Amplification; D008969:Molecular Sequence Data; D009231:Myxococcales / Q000235:genetics / Q000378:metabolism*; D010444:Peptide Elongation Factor Tu / Q000235:genetics*; D010448:Peptide Initiation Factors / Q000235:genetics*; D039881:Prokaryotic Initiation Factor-2; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D001426:Bacterial Proteins; D017931:DNA Primers; D010448:Peptide Initiation Factors; D039881:Prokaryotic Initiation Factor-2; D019204:GTP-Binding Proteins; D010444:Peptide Elongation Factor Tu