TY - JOUR
T1 - A two-phagemid system for the creation of non-phage displayed antibody libraries approaching one trillion members
AU - Ostermeier, Marc
AU - Benkovic, Stephen J.
N1 - Funding Information:
We would like to thank Dr D.R. Burton for providing plasmid pTC01 and Dr K.F Jensen for providing strains SØ4038 and SØ6355. This work was supported by a NIH postdoctoral fellowship (1 F32 GM18560-01) to M.O.
PY - 2000/4/3
Y1 - 2000/4/3
N2 - We have designed a two-phagemid system for the construction of very large non-phage displayed Fab antibody libraries in E. coli approaching 1012 members. The system can accommodate both periplasmic and cytoplasmic Fab expression and should prove useful for the direct selection of functional antibodies by genetic techniques. We successfully alleviate problems of Fab vector instability and report a set of improved 5' primers for the amplification of mouse Ig V(H) repertoires from mouse spleen. These primers have no more than one mismatch in the last 11 bases for >95% of mouse Ig V(H) genes and minimize the amount of N-terminal amino acid changes while maintaining the flexibility of periplasmic or cytoplasmic antibody expression in E. coli. Copyright (C) 2000 Elsevier Science B.V.
AB - We have designed a two-phagemid system for the construction of very large non-phage displayed Fab antibody libraries in E. coli approaching 1012 members. The system can accommodate both periplasmic and cytoplasmic Fab expression and should prove useful for the direct selection of functional antibodies by genetic techniques. We successfully alleviate problems of Fab vector instability and report a set of improved 5' primers for the amplification of mouse Ig V(H) repertoires from mouse spleen. These primers have no more than one mismatch in the last 11 bases for >95% of mouse Ig V(H) genes and minimize the amount of N-terminal amino acid changes while maintaining the flexibility of periplasmic or cytoplasmic antibody expression in E. coli. Copyright (C) 2000 Elsevier Science B.V.
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U2 - 10.1016/S0022-1759(99)00245-8
DO - 10.1016/S0022-1759(99)00245-8
M3 - Article
C2 - 10725461
AN - SCOPUS:0034599418
VL - 237
SP - 175
EP - 186
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
SN - 0022-1759
IS - 1-2
ER -