Tagging of Escherichia coli proteins with new cassettes allowing in vivo systematic fluorescent and luminescent detection, and purification from physiological expression levels.
We designed cassettes allowing the systematic fusion of fluorescent or luminescent proteins preceded by the calmodulin binding peptide tag to the C-terminus of Escherichia coli proteins. The chromosomal insertion, and thus physiological expression level of these fusions, permits the study of protein localization by fluorescent microscopy and protein quantification, in vivo and dynamically in diverse conditions. Furthermore, the calmodulin binding peptide tag allows standard detection, affinity purification, and co-purification experiments. These cassettes are therefore very valuable for the versatility of experiments they make available for a given strain, from biochemistry to dynamic and in vivo studies.
LISM, CNRS, Aix-Marseille University, Marseille, France.
Astrid
A
Wahl
Pierre
P
Hubert
James N
JN
Sturgis
Emmanuelle
E
Bouveret
D001483:Base Sequence; D002148:Calmodulin-Binding Proteins / Q000032:analysis* / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology* / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000032:analysis* / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D008164:Luminescent Proteins / Q000032:analysis* / Q000235:genetics; D008856:Microscopy, Fluorescence / Q000379:methods*; D010743:Phospholipids / Q000378:metabolism
D002148:Calmodulin-Binding Proteins; D029968:Escherichia coli Proteins; D008164:Luminescent Proteins; D010743:Phospholipids