Patent ID: 8603950
Filing Date: 2013-12-10
Classification: C07K,C40B

Abstract:
1. A method of preparing a library of antibody-encoding polynucleotides, which method comprises: (a) preparing a first library of nucleic acid sequences, wherein each nucleic acid sequence encodes a CDR domain of an antibody heavy chain variable region consisting of CDR3, (b) preparing a second library of nucleic acid sequences, each of which encodes a CDR domain of an antibody light chain variable region consisting of CDR3, (c) preparing a third library of nucleic acid sequences, wherein each nucleic acid sequence encodes a template heavy chain variable region domain, wherein the template heavy chain variable region domain comprises a CDR domain consisting of endogenous germline CDR1 and endogenous germline CDR2, (d) preparing a fourth library of nucleic acid sequences, wherein each nucleic acid sequence encodes a template light chain variable region domain, wherein the template light chain variable region domain comprises a CDR domain consisting of endogenous germline CDR1 and endogenous germline CDR2, (e) preparing a fifth library of nucleic acid sequences, wherein each nucleic acid sequence encodes an antibody constant region template, (f) operably linking the nucleic acid sequences of the libraries of (a), (c), and (e) in a first expression vector to produce a library of antibody heavy chain-encoding polynucleotides comprising the CDR1 and CDR2 of the third library and the CDR3 of the first library, (g) operably linking the nucleic acid sequences of the libraries of (b), (d), and (e) in a second expression vector to produce a library of antibody light chain-encoding polynucleotides comprising the CDR1 and CDR2 of the fourth library and the CDR3 of the second library, (h) transforming host cells with both the library of antibody heavy chain-encoding polynucleotides produced in step (f) and the library of antibody light chain-encoding polynucleotides produced in step (g), wherein the host cells express or can be induced to express Activation Induced Cytidine Deaminase (AID), and (i) expressing AID in the host cells, whereupon AID induces one or more mutations in the nucleic acid sequences of the library produced in step (h) and a library of antibody-encoding polynucleotides is produced.