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

Gene Therapy volume 30, pages 167–171 (2023)Cite this article

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Abstract

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.

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Fig. 1: Efficient electrotransfection of human B-cell lines and PBMCs.
Fig. 2: Efficient generation of TP53−/− RPMI8866 cells.

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Acknowledgements

This research was supported by grants from the GEFLUC, LNCC, the Presidium of RAS and the State program of fundamental scientific research of IDB RAS. We gratefully acknowledge the technical support of the imaging and cytometry platform of the Gustave Roussy Institute.

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Authors and Affiliations

  1. UMR 9018, CNRS, Univ. Paris-Sud, Université Paris Saclay, Institut Gustave Roussy, 94805, Villejuif, France

    Reynand Jay Canoy, Franck André, Anna Shmakova, Joëlle Wiels, Marc Lipinski, Yegor Vassetzky & Diego Germini

  2. Institute of Human Genetics, National Institutes of Health, University of the Philippines Manila, 1000, Manila, Philippines

    Reynand Jay Canoy

  3. Laboratory of molecular endocrinology, Institute of Experimental Cardiology, Federal State Budgetary Organization National Cardiology Research Center Ministry of Health of the Russian Federation, 121552, Moscow, Russia

    Anna Shmakova

  4. Koltzov Institute of Developmental Biology, RAS, 117334, Moscow, Russia

    Yegor Vassetzky

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  1. Reynand Jay Canoy
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  2. Franck André
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Correspondence to Yegor Vassetzky or Diego Germini.

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Canoy, R.J., André, F., Shmakova, A. et al. Easy and robust electrotransfection protocol for efficient ectopic gene expression and genome editing in human B cells. Gene Ther 30, 167–171 (2023). https://doi.org/10.1038/s41434-020-00194-x

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