An improved phage display vector for antibody repertoire cloning by construction of combinatorial libraries.
Phagemid pComb3 is a widely used vector for molecular cloning of the antibody repertoire and for production of phage display libraries. However, in practical use, the utilization of this vector has some drawbacks. In this work we describe the construction of pComb3/TIG, an improved, easily manipulated vector for the cloning and display of antibody fragment libraries on the surface of filamentous phage. The two small "stuffer" fragments at the cloning sites were replaced with long DNA fragments, for easier differentiation of the correctly cut forms of the vector. Moreover, in pComb3/TIG the fragment at the heavy-chain-fragment cloning site contains an acid phosphatase-encoding gene. This feature allows the easy distinction of the Escherichia coli cells containing the unmodified form of the phagemid instead of the heavy-chain fragment coding cDNA in a simple plate histochemical assay.
Burioni
R
R
Istituto di Microbiologia, Facoltà di Medicina, Università Cattolica del Sacro Cuore, Roma, Italy.
Plaisant
P
P
Delli Carri
V
V
Vannini
A
A
Spanu
T
T
Clementi
M
M
Fadda
G
G
Varaldo
P E
PE
D000906:Antibodies / Q000235:genetics*; D001435:Bacteriophages / Q000235:genetics*; D003001:Cloning, Molecular; D005822:Genetic Vectors; D006801:Humans
D000906:Antibodies