Work

Protocol for ecotropic pseudotyped lentiviral transduction of “murinized” human cells while decreasing risk of infection for laboratory personnel

Year: 2025

Type: article

Abstract: Lentiviral systems of transduction are valuable tools to stably express transgenes of interest in human cell lines. However, their use risks infection of the lab workers. Here, we present a protocol for lentiviral vector system pseudotyping with the murine ecotropic envelope. We detail steps for producing ecotropic pseudotyped lentiviral particles and transiently expressing the Slc7a1 receptor for murine leukemia virus in target human cells. We then describe procedures for puromycin selection and the characterization of stable cell lines for transgene expression. For complete details on the use and execution of this protocol, please refer to Iyer-Bierhoff et al. 1 • Steps for the production of ecotropic pseudotyped lentiviral particles • Instructions for “murinizing” human target cells with the membrane receptor Slc7a1 • Guide to lentiviral transduction of “murinized” target cells • Steps for puromycin selection and characterization of stable transductants Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Lentiviral systems of transduction are valuable tools to stably express transgenes of interest in human cell lines. However, their use risks infection of the lab workers. Here, we present a protocol for lentiviral vector system pseudotyping with the murine ecotropic envelope. We detail steps for producing ecotropic pseudotyped lentiviral particles and transiently expressing the Slc7a1 receptor for murine leukemia virus in target human cells. We then describe procedures for puromycin selection and the characterization of stable cell lines for transgene expression.

Source: STAR Protocols

DOI: 10.1016/j.xpro.2025.103708

Authors: Sandra L.K. Weber, Thorsten Heinzel, Aishwarya Iyer-Bierhoff

Institution: Friedrich Schiller University Jena

Language: English

Topic: Virus-based gene therapy research

Subfield: Genetics

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

SDG: Good health and well-being

Open Access status: gold

Estimated APC paid: $1,550