A general approach to detect protein expression in vivo using fluorescent puromycin conjugates.
Understanding the expression of known and unknown gene products represents one of the key challenges in the post-genomic world. Here, we have developed a new class of reagents to examine protein expression in vivo that does not require transfection, radiolabeling, or the prior choice of a candidate gene. To do this, we constructed a series of puromycin conjugates bearing various fluorescent and biotin moieties. These compounds are readily incorporated into expressed protein products in cell lysates in vitro and efficiently cross cell membranes to function in protein synthesis in vivo as indicated by flow cytometry, selective enrichment studies, and Western analysis. Overall, this work demonstrates that fluorescent-puromycin conjugates offer a general means to examine protein expression in vivo.
Starck
Shelley R
SR
Division of Chemistry and Chemical Engineering, California Institute of Technology, 1200 E. California Boulevard, Pasadena, California 91125, USA.
Green
Harry M
HM
Alberola-Ila
José
J
Roberts
Richard W
RW
D015153:Blotting, Western; D002460:Cell Line; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D011691:Puromycin / Q000737:chemistry*
D005456:Fluorescent Dyes; D011506:Proteins; D011691:Puromycin