2026-04-26T23:12:04Zhttps://repisalud.isciii.es/rest/oai/requestoai:repisalud.isciii.es:20.500.12105/261522025-12-18T12:53:53Zcom_20.500.12105_19604com_20.500.12105_2051col_20.500.12105_19605
Repisalud
author
Torres, Raul
author
García, Aida
author
Payá, Monica
author
Ramirez, Juan C
funder
Ministerio de Ciencia e Innovación (España)
2025-01-27T14:02:57Z
2025-01-27T14:02:57Z
2011
PLoS One. 2011 May 23;6(5):e19794.
https://hdl.handle.net/20.500.12105/26152
21625434
PLoS One
Recombinase mediated cassette exchange (RMCE) is a two-step process leading to genetic modification in a specific genomic target sequence. The process involves insertion of a docking genetic cassette in the genome followed by DNA transfer of a second cassette flanked by compatible recombination signals and expression of the recombinase. Major technical drawbacks are cell viability upon transfection, toxicity of the enzyme, and the ability to target efficiently cell types of different origins. To overcome such drawbacks, we developed an RMCE assay that uses an integrase-deficient lentivirus (IDLV) vector in the second step combined with promoterless trapping of double selectable markers. Additionally, recombinase expression is self-limiting as a result of the exchangeable reaction, thus avoiding toxicity. Our approach provides proof-of-principle of a simple and novel strategy with expected wide applicability modelled on a human cell line with randomly integrated copies of a genetic landing pad. This strategy does not present foreseeable limitations for application to other cell systems modified by homologous recombination. Safety, efficiency, and simplicity are the major advantages of our system, which can be applied in low-to-medium throughput strategies for screening of cDNAs, non-coding RNAs during functional genomic studies, and drug screening.
eng
Non-integrative lentivirus drives high-frequency cre-mediated cassette exchange in human cells.
research article
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URL
https://repisalud.isciii.es/bitstreams/3a67fa42-c515-4c6f-a08f-4b75cb151472/download
File
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bc3dfc1d8eb70a99684a808d67565d1c
665375
application/pdf
Non-Integrative Lentivirus_PLoS One_2011.pdf