Highly efficient multiplex genetic engineering of porcine primary fetal fibroblasts.

Benjamin Klapholz, Heather Levy, Ramesh Kumbha, Nora Hosny, Michael E D'Angelo, Bernhard J Hering, Christopher Burlak

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Article in Surgery open science, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

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5 · Who and what money

Authors and funding

7 authors.

Benjamin KlapholzHorizon Discovery, 8100 Cambridge Research Park, Waterbeach, Cambridge CB25 9TL, UK.

Heather LevySchulze Diabetes Institute, Department of Surgery, University of Minnesota Medical School, Minneapolis, MN, USA.

Ramesh KumbhaSchulze Diabetes Institute, Department of Surgery, University of Minnesota Medical School, Minneapolis, MN, USA.

Nora HosnyDepartment of Surgery, Schulze Diabetes Institute, University of Minnesota School of Medicine, Minneapolis, MN, USA.

Michael E D'AngeloHorizon Discovery, 8100 Cambridge Research Park, Waterbeach, Cambridge CB25 9TL, UK.

Bernhard J HeringSchulze Diabetes Institute, Department of Surgery, University of Minnesota Medical School, Minneapolis, MN, USA.

Christopher BurlakSchulze Diabetes Institute, Department of Surgery, University of Minnesota Medical School, Minneapolis, MN, USA.

Funding

No grant is acknowledged in the PubMed record.

6 · The paper itself

Abstract

backgroundGenetically engineered porcine donors are a potential solution for the shortage of human organs for transplantation. Incompatibilities between humans and porcine donors are largely due to carbohydrate xenoantigens on the surface of porcine cells, provoking an immune response which leads to xenograft rejection. MATERIALS AND

methodsMultiplex genetic knockout of GGTA1, β4GalNT2, and CMAH is predicted to increase the rate of xenograft survival, as described previously for GGTA1. In this study, the clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated protein 9 system was used to target genes relevant to xenotransplantation, and a method for highly efficient editing of multiple genes in primary porcine fibroblasts was described.

resultsEditing efficiencies greater than 85% were achieved for knockout of GGTA1, β4GalNT2, and CMAH.

conclusionThe high-efficiency protocol presented here reduces scale and cost while accelerating the production of genetically engineered primary porcine fibroblast cells for in vitro studies and the production of animal models.

Identifiers

PMID33937740

DOI10.1016/j.sopen.2020.11.003

PMCPMC8074785

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