PRJNA450158
Homo sapiens isolate:Ramos (RA 1) B lymphocyte Genome sequencing and assembly
A homology-directed repair (HDR) strategy was used to graft donor DNA containing the VDJ gene from a broadly neutralizing HIV antibody (PG9), between the 5'' most V-gene promoter (V781) and the J6 splice site in the human Ramos (RA 1) B cell line IGHV locus. Engineered cells expressing PG9 HC using the Ramos µ constant gene could be enriched using FACS with HIV envelope probes which recognized the PG9 HC/Ramos ?LC cell surface expressed IgM. This was compared with a second HDR strategy directly replacing the expressed Ramos VDJ variable region which used the V4-34 and J6 genes. Engineered HCs retained the ability to class switch and undergo somatic hypermutation which generated variants with higher HIV neutralization breadth and potency.