Abstract
Recombinant monoclonal antibodies can be established by displaying single-chain variable fragment (scFv) antibody libraries on phages and then biopanning against the target. For constructing superior scFv libraries, antibody light-chain variable region (VL) and heavy-chain variable region (VH) fragments must be assembled into scFvs without loss of diversity. A high-quality scFv library is a prerequisite for obtaining strong binders from the scFv library. However, the technical challenges associated with the construction of a diverse library have been the bottleneck in the establishment of recombinant antibodies through biopanning. Here, we describe a simple and efficient method for assembling VL and VH fragments through the concerted action of λ-exonuclease and Bst DNA polymerase. We successfully used this method to construct a diverse chicken scFv library.
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Abbreviations
- scFv:
-
Single-chain variable fragment
- VL :
-
Light-chain variable region
- VH:
-
Heavy-chain variable region
- CDR:
-
Complementarity-determining region
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Kato, M., Hanyu, Y. (2017). Enzymatic Assembly for scFv Library Construction. In: Tiller, T. (eds) Synthetic Antibodies. Methods in Molecular Biology, vol 1575. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6857-2_3
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DOI: https://doi.org/10.1007/978-1-4939-6857-2_3
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