Easy and robust electrotransfection protocol for efficient ectopic gene expression and genome editing in human B cells

• Published in: Gene therapy Vol. 30; no. 1-2; pp. 167 - 171
• Main Authors: Canoy, Reynand Jay, André, Franck, Shmakova, Anna, Wiels, Joëlle, Lipinski, Marc, Vassetzky, Yegor, Germini, Diego
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01-02-2023; Nature Publishing Group
• Subjects: 42/109; 42/41; 631/1647/2300/1851; 631/67/1990/291/1621/1915; B-cell lymphoma; Biomedical and Life Sciences; Biomedicine; Biotechnology; Brief Communication; Cardiology; Cell Biology; Cell Line; Cells; Ectopic Gene Expression; Efficiency; Electric fields; Gene Editing - methods; Gene Expression; Gene silencing; Gene Therapy; Genome editing; Genomes; Human Genetics; Humans; Laboratories; Leukemia; Life Sciences; Lymphocytes B; Lymphoma; Nanotechnology; p53 Protein; Plasmids; Protocol; Transfection; Tumor cell lines
• ISSN: 0969-7128; 1476-5462
• DOI: 10.1038/s41434-020-00194-x

B-cell lines and primary PBMCs are notoriously hard to transfect, thus making genome editing, ectopic gene expression, or gene silencing experiments particularly tedious. Here we propose a novel efficient and reproducible protocol for electrotransfection of lymphoblastoid, B-cell lymphoma, leukemia cell lines, and B cells from PBMCs. The proposed protocol requires neither costly equipment nor expensive reagents; it can be used with small or large plasmids. Transfection and viability rates of about 79% and 58%, respectively, have been routinely achieved by optimizing the salt concentration in the electrotransfection medium and the amount of plasmid used. A validation of the protocol was obtained
via
the generation of a
TP53
−/
−
RPMI8866 lymphoblastoid cell line which should prove useful in future hematological and blood cancer studies.