Large-scale production of dsRNA and siRNA pools for RNA interference utilizing bacteriophage varphi6 RNA-dependent RNA polymerase. Export

@article{citeulike:1066564, abstract = {The discovery of RNA interference (RNAi) has revolutionized biological research and has a huge potential for therapy. Since small double-stranded RNAs (dsRNAs) are required for various RNAi applications, there is a need for cost-effective methods for producing large quantities of high-quality dsRNA. We present two novel, flexible virus-based systems for the efficient production of dsRNA: (1) an in vitro system utilizing the combination of T7 RNA polymerase and RNA-dependent RNA polymerase (RdRP) of bacteriophage varphi6 to generate dsRNA molecules of practically unlimited length, and (2) an in vivo RNA replication system based on carrier state bacterial cells containing the varphi6 polymerase complex to produce virtually unlimited amounts of dsRNA of up to 4.0 kb. We show that pools of small interfering RNAs (siRNAs) derived from dsRNA produced by these systems significantly decreased the expression of a transgene (eGFP) in HeLa cells and blocked endogenous pro-apoptotic BAX expression and subsequent cell death in cultured sympathetic neurons.}, address = {1Institute of Biotechnology and Department of Biological and Environmental Sciences, University of Helsinki, Biocenter 2, FIN-00014 University of Helsinki, Helsinki, Finland.}, author = {Aalto, Antti P P. and Sarin, L Peter P. and van Dijk, Alberdina A A. and Saarma, Mart and Poranen, Minna M M. and Arum\"{a}e, Urmas and Bamford, Dennis H H.}, citeulike-article-id = {1066564}, citeulike-linkout-0 = {http://dx.doi.org/10.1261/rna.348307}, citeulike-linkout-1 = {http://www.rnajournal.org/cgi/content/abstract/13/3/422}, citeulike-linkout-2 = {http://view.ncbi.nlm.nih.gov/pubmed/17237359}, citeulike-linkout-3 = {http://www.hubmed.org/display.cgi?uids=17237359}, day = {19}, doi = {10.1261/rna.348307}, issn = {1355-8382}, journal = {RNA}, month = {January}, posted-at = {2007-10-25 16:30:01}, priority = {2}, title = {Large-scale production of dsRNA and siRNA pools for RNA interference utilizing bacteriophage {varphi}6 RNA-dependent RNA polymerase.}, url = {http://dx.doi.org/10.1261/rna.348307}, year = {2007} }

TY - JOUR ID - citeulike:1066564 L3 - citeulike-article-id:1066564 N2 - The discovery of RNA interference (RNAi) has revolutionized biological research and has a huge potential for therapy. Since small double-stranded RNAs (dsRNAs) are required for various RNAi applications, there is a need for cost-effective methods for producing large quantities of high-quality dsRNA. We present two novel, flexible virus-based systems for the efficient production of dsRNA: (1) an in vitro system utilizing the combination of T7 RNA polymerase and RNA-dependent RNA polymerase (RdRP) of bacteriophage varphi6 to generate dsRNA molecules of practically unlimited length, and (2) an in vivo RNA replication system based on carrier state bacterial cells containing the varphi6 polymerase complex to produce virtually unlimited amounts of dsRNA of up to 4.0 kb. We show that pools of small interfering RNAs (siRNAs) derived from dsRNA produced by these systems significantly decreased the expression of a transgene (eGFP) in HeLa cells and blocked endogenous pro-apoptotic BAX expression and subsequent cell death in cultured sympathetic neurons. JF - RNA SN - 1355-8382 AD - 1Institute of Biotechnology and Department of Biological and Environmental Sciences, University of Helsinki, Biocenter 2, FIN-00014 University of Helsinki, Helsinki, Finland. TI - Large-scale production of dsRNA and siRNA pools for RNA interference utilizing bacteriophage {varphi}6 RNA-dependent RNA polymerase. AU - Aalto, Antti P AU - Sarin, L Peter AU - van Dijk, Alberdina A AU - Saarma, Mart AU - Poranen, Minna M AU - Arumäe, Urmas AU - Bamford, Dennis H PY - 2007/01/19/ UR - http://dx.doi.org/10.1261/rna.348307 ER -