A novel vector for positive selection of inserts harboring an open reading frame by translational coupling.
We have developed a novel vector pTCS, as a tool for efficient selection of open reading frame (ORF)-containing inserts. In pTCS clones containing an insert with an ORF a downstream marker gene (immE3, conferring resistance to colicin) is activated via translational coupling with the insert, and transformed cells can then be selected by exposure to colicin E3. Our method differs from previous methods in that the marker gene is activated without protein fusion, and that selection occurs irrespective of the reading frame of the insert.
Ohashi-Kunihiro
Sumiko
S
Research Institute for Cell Engineering, National Institute of Advanced Industrial Science and Technology (AIST), Higashi, Tsukuba, Ibaraki, Japan.
Yohda
Masafumi
M
Masaki
Haruhiko
H
Machida
Masayuki
M
D003086:Bacteriocin Plasmids / Q000235:genetics; D001483:Base Sequence; D003087:Colicins / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005819:Genetic Markers; D005822:Genetic Vectors / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000379:methods*; D016366:Open Reading Frames / Q000235:genetics*; D014176:Protein Biosynthesis
D003087:Colicins; D005819:Genetic Markers