A cloning vector for efficient generation of cholera toxin B gene fusions for epitope screening.
Gene fusion proteins with epitopes attached to the amino end of cholera toxin B subunit (CTB) are useful to raise immunological responses. We describe a cloning vector, designated pCTBtet, carrying a tetracycline resistance gene (TetR) between the leader peptide and mature CTB. Removal of TetR to insert oligonucleotides encoding fusion epitopes allowed for screening of tetracycline-sensitive clones. Restoration of the correct CTB reading phase was subsequently used to choose gene fusion candidate colonies. The use of pCTBtet permitted the rapid construction of 8 fusion proteins carrying 9-24 aa from Salmonella typhi OmpC and 6 hybrids with 7-31 aa from Escherichia coli colonization factor CFAI.
Secundino
I
I
Unidad de Investigación Médica en Inmunoquímica, Hospital de Especialidades, Centro Médico Nacional Siglo XXI, IMSS. México, D.F.
Paniagua-Solís
J
J
Isibasi
A
A
Sanchez
J
J
D000595:Amino Acid Sequence; D019677:Artificial Gene Fusion; D001483:Base Sequence; D015153:Blotting, Western; D002772:Cholera Toxin / Q000737:chemistry / Q000235:genetics*; D003001:Cloning, Molecular; D017931:DNA Primers; D004587:Electrophoresis, Agar Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D008969:Molecular Sequence Data; D010957:Plasmids; D013753:Tetracycline Resistance / Q000235:genetics
D017931:DNA Primers; D000939:Epitopes; D002772:Cholera Toxin