Diversity analysis of an immune VHH (Variable Heavy domain from Heavy chain only antibodies) library before and after its integration in E. coli chromosome.

In this study we aim to determine the diversity of a gene library of VHHs before and after its integration in the E. coli chromosome. The final objective is to demonstrate that the library diversity is maintained after the integration process. The original library was composed of a pool of bacterial plasmids containing the VHHs which finally were integrated into the E. coli chromosome at a specific gene locus. The diversity of the non-integrated library was analyzed by Miseq pair-end of VHH genes amplified from the plasmid collection while the diversity of the integrated library was analyzed from VHH amplicons obtained from genomic DNA of the integrated bacterial pool. The relevance of this project lies in the need to preserve genetic diversity when working with large gene libraries. The library used in this study contained ca. 13 million VHHs genes from Llama (Lama glama) obtained after immunization with A431 human tumor cell cell line. Around 500000 sequences representing ca. 3.8 % of the sequences from the original library were analyzed.