Diversity analysis of a naive 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 naive 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 4 pools 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 4 pools of plasmids while the diversity of the integrated library was analyzed from 2 VHH amplicons obtained from genomic DNA of the integrated bacterial pools. 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. 1000 million VHHs genes from naive (non-immunized) dromedaries (Camelus dromedarius). Around 2 x 106 sequences from the non-integrated plasmid library and 1 million sequences from the integrated library were analyzed representing ca. 0.1-0.2 % of the sequences from the original library.