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A cross-species whole genome siRNA screen in suspension-cultured Chinese hamster ovary cells identifies novel engineering targets.
Klanert, Gerald; Fernandez, Daniel J; Weinguny, Marcus; Eisenhut, Peter; Bühler, Eugen; Melcher, Michael; Titus, Steven A; Diendorfer, Andreas B; Gludovacz, Elisabeth; Jadhav, Vaibhav; Xiao, Su; Stern, Beate; Lal, Madhu; Shiloach, Joseph; Borth, Nicole.
Afiliación
  • Klanert G; Austrian Centre of Industrial Biotechnology, Graz, Austria.
  • Fernandez DJ; University of Natural Resources and Life Sciences, Vienna, Austria.
  • Weinguny M; Division of Preclinical Innovation, NCATS, NIH, Rockville, MD, USA.
  • Eisenhut P; Austrian Centre of Industrial Biotechnology, Graz, Austria.
  • Bühler E; University of Natural Resources and Life Sciences, Vienna, Austria.
  • Melcher M; Austrian Centre of Industrial Biotechnology, Graz, Austria.
  • Titus SA; University of Natural Resources and Life Sciences, Vienna, Austria.
  • Diendorfer AB; Division of Preclinical Innovation, NCATS, NIH, Rockville, MD, USA.
  • Gludovacz E; Austrian Centre of Industrial Biotechnology, Graz, Austria.
  • Jadhav V; University of Natural Resources and Life Sciences, Vienna, Austria.
  • Xiao S; Division of Preclinical Innovation, NCATS, NIH, Rockville, MD, USA.
  • Stern B; Austrian Centre of Industrial Biotechnology, Graz, Austria.
  • Lal M; University of Natural Resources and Life Sciences, Vienna, Austria.
  • Shiloach J; Medical University of Vienna, Vienna, Austria.
  • Borth N; Austrian Centre of Industrial Biotechnology, Graz, Austria.
Sci Rep ; 9(1): 8689, 2019 06 18.
Article en En | MEDLINE | ID: mdl-31213643
High-throughput siRNA screens were only recently applied to cell factories to identify novel engineering targets which are able to boost cells towards desired phenotypes. While siRNA libraries exist for model organisms such as mice, no CHO-specific library is publicly available, hindering the application of this technique to CHO cells. The optimization of these cells is of special interest, as they are the main host for the production of therapeutic proteins. Here, we performed a cross-species approach by applying a mouse whole-genome siRNA library to CHO cells, optimized the protocol for suspension cultured cells, as this is the industrial practice for CHO cells, and developed an in silico method to identify functioning siRNAs, which also revealed the limitations of using cross-species libraries. With this method, we were able to identify several genes that, upon knockdown, enhanced the total productivity in the primary screen. A second screen validated two of these genes, Rad21 and Chd4, whose knockdown was tested in additional CHO cell lines, confirming the induced high productivity phenotype, but also demonstrating the cell line/clone specificity of engineering effects.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Biblioteca de Genes / Genoma / ARN Interferente Pequeño / Ensayos Analíticos de Alto Rendimiento Tipo de estudio: Guideline / Prognostic_studies Límite: Animals / Humans Idioma: En Revista: Sci Rep Año: 2019 Tipo del documento: Article País de afiliación: Austria Pais de publicación: Reino Unido
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Biblioteca de Genes / Genoma / ARN Interferente Pequeño / Ensayos Analíticos de Alto Rendimiento Tipo de estudio: Guideline / Prognostic_studies Límite: Animals / Humans Idioma: En Revista: Sci Rep Año: 2019 Tipo del documento: Article País de afiliación: Austria Pais de publicación: Reino Unido