Abstract
RNA interference has become an indispensable tool for loss-of-function studies across eukaryotes. By enabling stable and reversible gene silencing, shRNAs provide a means to study long-term phenotypes, perform pool-based forward genetic screens and examine the consequences of temporary target inhibition in vivo. However, efficient implementation in vertebrate systems has been hindered by technical difficulties affecting potency and specificity. Focusing on these issues, we analyse current strategies to obtain maximal knockdown with minimal off-target effects.
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References
Hopkins, A. L. & Groom, C. R. The druggable genome. Nat. Rev. Drug Discov. 1, 727–730 (2002).
Bartel, D. P. MicroRNAs: Genomics, biogenesis, mechanism, and function. Cell 116, 281–297 (2004).
Hannon, G. J. RNA interference. Nature 418, 244–251 (2002).
Bartel, D. P. MicroRNAs: Target recognition and regulatory functions. Cell 136, 215–233 (2009).
Filipowicz, W., Bhattacharyya, S. N. & Sonenberg, N. Mechanisms of post-transcriptional regulation by microRNAs: are the answers in sight? Nat. Rev. Genet. 9, 102–114 (2008).
Elbashir, S. M. et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411, 494–498 (2001).
Brummelkamp, T. R., Bernards, R. & Agami, R. A system for stable expression of short interfering RNAs in mammalian cells. Science 296, 550–553 (2002).
Paddison, P. J., Caudy, A. A., Bernstein, E., Hannon, G. J. & Conklin, D. S. Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells. Genes Dev. 16, 948–958 (2002).
McManus, M. T., Petersen, C. P., Haines, B. B., Chen, J. & Sharp, P. A. Gene silencing using micro-RNA designed hairpins. RNA 8, 842–850 (2002).
Zeng, Y., Wagner, E. J. & Cullen, B. R. Both natural and designed microRNAs can inhibit the expression of cognate mRNAs when expressed in human cells. Mol. Cell 9, 1327–1333 (2002).
Silva, J. M. et al. Second-generation shRNA libraries covering the mouse and human genome. Nat. Genet. 37, 1281–1288 (2005).
Chung, K. H. et al. Polycistronic RNA polymerase II expression vectors for RNA interference based on BIC/miR-155. Nucleic Acids Res. 34, e53 (2006).
Ely, A., Naidoo, T., Mufamadi, S., Crowther, C. & Arbuthnot, P. Expressed anti-HBV primary microRNA shuttles inhibit viral replication efficiently in vitro and in vivo. Mol. Ther. 16, 1105–1112 (2008).
Aagaard, L. A. et al. Engineering and optimization of the miR-106b cluster for ectopic expression of multiplexed anti-HIV RNAs. Gene Ther. 15, 1536–1549 (2008).
Liu, Y. P., Haasnoot, J., ter Brake, O., Berkhout, B. & Konstantinova, P. Inhibition of HIV-1 by multiple siRNAs expressed from a single microRNA polycistron. Nucleic Acids Res. 36, 2811–2824 (2008).
McBride, J. L. et al. Artificial miRNAs mitigate shRNA-mediated toxicity in the brain: implications for the therapeutic development of RNAi. Proc. Natl Acad. Sci. USA 105, 5868–5873 (2008).
Boudreau, R. L., Martins, I. & Davidson, B. L. Artificial microRNAs as siRNA shuttles: improved safety as compared to shRNAs in vitro and in vivo. Mol. Ther. 17, 169–175 (2008).
Castanotto, D. et al. Combinatorial delivery of small interfering RNAs reduces RNAi efficacy by selective incorporation into RISC. Nucleic Acids Res. 35, 5154–5164, (2007).
Grimm, D. et al. Fatality in mice due to oversaturation of cellular microRNA/short hairpin RNA pathways. Nature 441, 537–541 (2006).
Dickins, R. A. et al. Probing tumor phenotypes using stable and regulated synthetic microRNA precursors. Nat. Genet. 37, 1289–1295 (2005).
Stegmeier, F., Hu, G., Rickles, R. J., Hannon, G. J. & Elledge, S. J. A lentiviral microRNA-based system for single-copy polymerase II-regulated RNA interference in mammalian cells. Proc. Natl Acad. Sci. USA 102, 13212–13217 (2005).
Jackson, A. L. et al. Expression profiling reveals off-target gene regulation by RNAi. Nat. Biotechnol. 21, 635–637 (2003).
Birmingham, A. et al. 3′ UTR seed matches, but not overall identity, are associated with RNAi off-targets. Nat. Methods 3, 199–204 (2006).
Fellmann, C. et al. Functional identification of optimized RNAi triggers using a massively parallel sensor assay. Mol. Cell 41, 733–746 (2011).
Reynolds, A. et al. Rational siRNA design for RNA interference. Nat. Biotechnol. 22, 326–330 (2004).
Schwarz, D. S. et al. Asymmetry in the assembly of the RNAi enzyme complex. Cell 115, 199–208 (2003).
Khvorova, A., Reynolds, A. & Jayasena, S. D. Functional siRNAs and miRNAs exhibit strand bias. Cell 115, 209–216 (2003).
Huesken, D. et al. Design of a genome-wide siRNA library using an artificial neural network. Nat. Biotechnol. 23, 995–1001 (2005).
Vert, J. P., Foveau, N., Lajaunie, C. & Vandenbrouck, Y. An accurate and interpretable model for siRNA efficacy prediction. BMC Bioinformatics 7, 520 (2006).
Birmingham, A. et al. A protocol for designing siRNAs with high functionality and specificity. Nat. Protoc. 2, 2068–2078 (2007).
Pei, Y. & Tuschl, T. On the art of identifying effective and specific siRNAs. Nat. Methods 3, 670–676 (2006).
Taxman, D. J. et al. Criteria for effective design, construction, and gene knockdown by shRNA vectors. BMC Biotechnol. 6, 7 (2006).
Bassik, M. C. et al. Rapid creation and quantitative monitoring of high coverage shRNA libraries. Nat. Methods 6, 443–445 (2009).
Du, Q., Thonberg, H., Zhang, H. Y., Wahlestedt, C. & Liang, Z. Validating siRNA using a reporter made from synthetic DNA oligonucleotides. Biochem. Biophys. Res. Commun. 325, 243–249 (2004).
Kumar, R., Conklin, D. S. & Mittal, V. High-throughput selection of effective RNAi probes for gene silencing. Genome Res. 13, 2333–2340 (2003).
Mayr, C. & Bartel, D. P. Widespread shortening of 3′UTRs by alternative cleavage and polyadenylation activates oncogenes in cancer cells. Cell 138, 673–684 (2009).
Sandberg, R., Neilson, J. R., Sarma, A., Sharp, P. A. & Burge, C. B. Proliferating cells express mRNAs with shortened 3′ untranslated regions and fewer microRNA target sites. Science 320, 1643–1647 (2008).
Tan, X. et al. Tiling genomes of pathogenic viruses identifies potent antiviral shRNAs and reveals a role for secondary structure in shRNA efficacy. Proc. Natl Acad. Sci. USA 109, 869–874 (2012).
Naito, Y. & Ui-Tei, K. siRNA design software for a target gene-specific RNA interference. Front. Genet. 3, 102 (2012).
Matveeva, O. V., Nazipova, N. N., Ogurtsov, A. Y. & Shabalina, S. A. Optimized models for design of efficient miR30-based shRNAs. Front. Genet. 3, 163 (2012).
Kaelin, W. G. Jr. Use and abuse of RNAi to study mammalian gene function. Science 337, 421–422 (2012).
Jackson, A. L. & Linsley, P. S. Recognizing and avoiding siRNA off-target effects for target identification and therapeutic application. Nat. Rev. Drug Discov. 9, 57–67 (2010).
Grimson, A. et al. MicroRNA targeting specificity in mammals: determinants beyond seed pairing. Mol. Cell 27, 91–105 (2007).
Semizarov, D. et al. Specificity of short interfering RNA determined through gene expression signatures. Proc. Natl Acad. Sci. USA 100, 6347–6352 (2003).
Yi, R., Doehle, B. P., Qin, Y., Macara, I. G. & Cullen, B. R. Overexpression of exportin 5 enhances RNA interference mediated by short hairpin RNAs and microRNAs. RNA 11, 220–226 (2005).
Khan, A. A. et al. Transfection of small RNAs globally perturbs gene regulation by endogenous microRNAs. Nat. Biotechnol. 27, 549–555 (2009).
Premsrirut, P. K. et al. A rapid and scalable system for studying gene function in mice using conditional RNA interference. Cell 145, 145–158 (2011).
Gu, S. et al. The loop position of shRNAs and pre-miRNAs is critical for the accuracy of Dicer processing in vivo. Cell 151, 900–911 (2012).
Kawahara, Y. et al. Redirection of silencing targets by adenosine-to-inosine editing of miRNAs. Science 315, 1137–1140 (2007).
Moffat, J. & Sabatini, D. M. Building mammalian signalling pathways with RNAi screens. Nat. Rev. Mol. Cell Biol. 7, 177–187 (2006).
Ngo, V. N. et al. A loss-of-function RNA interference screen for molecular targets in cancer. Nature 441, 106–110 (2006).
Westbrook, T. F. et al. A genetic screen for candidate tumor suppressors identifies REST. Cell 121, 837–848 (2005).
Solimini, N. L. et al. Recurrent hemizygous deletions in cancers may optimize proliferative potential. Science 337, 104–109 (2012).
Meacham, C. E., Ho, E. E., Dubrovsky, E., Gertler, F. B. & Hemann, M. T. In vivo RNAi screening identifies regulators of actin dynamics as key determinants of lymphoma progression. Nat. Genet. 41, 1133–1137 (2009).
Prahallad, A. et al. Unresponsiveness of colon cancer to BRAF(V600E) inhibition through feedback activation of EGFR. Nature 483, 100–103 (2012).
Cheung, H. W. et al. Systematic investigation of genetic vulnerabilities across cancer cell lines reveals lineage-specific dependencies in ovarian cancer. Proc. Natl Acad. Sci. USA 108, 12372–12377 (2011).
Conrad, C. & Gerlich, D. W. Automated microscopy for high-content RNAi screening. J. Cell Biol. 188, 453–461 (2010).
Neumann, B. et al. Phenotypic profiling of the human genome by time-lapse microscopy reveals cell division genes. Nature 464, 721–727 (2010).
Sigoillot, F. D. et al. A bioinformatics method identifies prominent off-targeted transcripts in RNAi screens. Nat. Methods 9, 363–366 (2012).
Shao, D. D. et al. ATARiS: computational quantification of gene suppression phenotypes from multisample RNAi screens. Genome Res. 23, 665–678 (2013).
Schmidt, E. E. et al. GenomeRNAi: a database for cell-based and in vivo RNAi phenotypes, 2013 update. Nucleic Acids Res. 41, D1021–D1026 (2013).
Bassik, M. C. et al. A systematic mammalian genetic interaction map reveals pathways underlying ricin susceptibility. Cell 152, 909–922 (2013).
Frese, K. K. & Tuveson, D. A. Maximizing mouse cancer models. Nat. Rev. Cancer 7, 645–658 (2007).
Schmitt, C. A., Rosenthal, C. T. & Lowe, S. W. Genetic analysis of chemoresistance in primary murine lymphomas. Nat. Med. 6, 1029–1035 (2000).
Zuber, J. et al. Mouse models of human AML accurately predict chemotherapy response. Genes Dev. 23, 877–889 (2009).
Zuber, J. et al. An integrated approach to dissecting oncogene addiction implicates a Myb-coordinated self-renewal program as essential for leukemia maintenance. Genes Dev. 25, 1628–1640 (2011).
Beard, C., Hochedlinger, K., Plath, K., Wutz, A. & Jaenisch, R. Efficient method to generate single-copy transgenic mice by site-specific integration in embryonic stem cells. Genesis 44, 23–28 (2006).
Gossen, M. & Bujard, H. Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc. Natl Acad. Sci. USA 89, 5547–5551 (1992).
Gossen, M. et al. Transcriptional activation by tetracyclines in mammalian cells. Science 268, 1766–1769 (1995).
Dickins, R. A. et al. Tissue-specific and reversible RNA interference in transgenic mice. Nat Genet 39, 914–921 (2007).
Hitz, C., Steuber-Buchberger, P., Delic, S., Wurst, W. & Kuhn, R. Generation of shRNA transgenic mice. Methods Mol. Biol. 530, 101–129 (2009).
Peng, S., York, J. P. & Zhang, P. A transgenic approach for RNA interference-based genetic screening in mice. Proc. Natl Acad. Sci. USA 103, 2252–2256 (2006).
McJunkin, K. et al. Reversible suppression of an essential gene in adult mice using transgenic RNA interference. Proc. Natl Acad. Sci. USA 108, 7113–7118 (2011).
Dow, L. E. & Lowe, S. W. Life in the fast lane: mammalian disease models in the genomics era. Cell 148, 1099–1109 (2012).
Lin, C. J. et al. Targeting synthetic lethal interactions between Myc and the eIF4F complex impedes tumorigenesis. Cell Rep. 1, 325–333 (2012).
Zuber, J. et al. RNAi screen identifies Brd4 as a therapeutic target in acute myeloid leukaemia. Nature 478, 524–528 (2011).
Hemann, M. T. et al. An epi-allelic series of p53 hypomorphs created by stable RNAi produces distinct tumor phenotypes in vivo. Nat. Genet. 33, 396–400 (2003).
Zender, L. et al. An oncogenomics-based in vivo RNAi screen identifies tumor suppressors in liver cancer. Cell 135, 852–864 (2008).
Bric, A. et al. Functional identification of tumor-suppressor genes through an in vivo RNA interference screen in a mouse lymphoma model. Cancer Cell 16, 324–335 (2009).
Scuoppo, C. et al. A tumour suppressor network relying on the polyamine-hypusine axis. Nature 487, 244–248 (2012).
Xue, W. et al. A cluster of cooperating tumor-suppressor gene candidates in chromosomal deletions. Proc. Natl Acad. Sci. USA 109, 8212–8217 (2012).
Possemato, R. et al. Functional genomics reveal that the serine synthesis pathway is essential in breast cancer. Nature 476, 346–350 (2011).
Miller, P. G. et al. In vivo RNAi screening identifies a leukemia-specific dependence on integrin beta 3 signaling. Cancer Cell 24, 45–58 (2013).
Wuestefeld, T. et al. A direct in vivo RNAi screen identifies MKK4 as a key regulator of liver regeneration. Cell 153, 389–401 (2013).
Zuber, J. et al. Toolkit for evaluating genes required for proliferation and survival using tetracycline-regulated RNAi. Nat. Biotechnol. 29, 79–83 (2011).
Okamura, K., Ishizuka, A., Siomi, H. & Siomi, M. C. Distinct roles for Argonaute proteins in small RNA-directed RNA cleavage pathways. Genes Dev. 18, 1655–1666 (2004).
Forstemann, K., Horwich, M. D., Wee, L., Tomari, Y. & Zamore, P. D. Drosophila microRNAs are sorted into functionally distinct argonaute complexes after production by dicer-1. Cell 130, 287–297 (2007).
Betancur, J. G., Yoda, M. & Tomari, Y. miRNA-like duplexes as RNAi triggers with improved specificity. Front. Genet. 3, 127 (2012).
Cleary, M. A. et al. Production of complex nucleic acid libraries using highly parallel in situ oligonucleotide synthesis. Nat. Methods 1, 241–248 (2004).
Loewer, S. et al. Large intergenic non-coding RNA-RoR modulates reprogramming of human induced pluripotent stem cells. Nat. Genet. 42, 1113–1117 (2010).
Wang, Y. et al. Endogenous miRNA sponge lincRNA-RoR regulates Oct4, Nanog, and Sox2 in human embryonic stem cell self-renewal. Dev. Cell 25, 69–80 (2013).
Castel, S. E. & Martienssen, R. A. RNA interference in the nucleus: roles for small RNAs in transcription, epigenetics and beyond. Nat. Rev. Genet. 14, 100–112 (2013).
Fu, Y. et al. High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells. Nat. Biotechnol. 31, 822–826 (2013).
Napoli, C., Lemieux, C. & Jorgensen, R. Introduction of a chimeric chalcone synthase gene into petunia results in reversible co-suppression of homologous genes in trans. Plant Cell 2, 279–289 (1990).
van der Krol, A. R., Mur, L. A., Beld, M., Mol, J. N. & Stuitje, A. R. Flavonoid genes in petunia: addition of a limited number of gene copies may lead to a suppression of gene expression. Plant Cell 2, 291–299 (1990).
Lee, R. C., Feinbaum, R. L. & Ambros, V. The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 75, 843–854 (1993).
Wightman, B., Ha, I. & Ruvkun, G. Posttranscriptional regulation of the heterochronic gene lin-14 by lin-4 mediates temporal pattern formation in C. elegans. Cell 75, 855–862 (1993).
Britten, R. J. & Davidson, E. H. Gene regulation for higher cells: a theory. Science 165, 349–357 (1969).
Jacob, F. & Monod, J. Genetic regulatory mechanisms in the synthesis of proteins. J. Mol. Biol. 3, 318–356 (1961).
Lagos-Quintana, M., Rauhut, R., Lendeckel, W. & Tuschl, T. Identification of novel genes coding for small expressed RNAs. Science 294, 853–858 (2001).
Lau, N. C., Lim, L. P., Weinstein, E. G. & Bartel, D. P. An abundant class of tiny RNAs with probable regulatory roles in Caenorhabditis elegans. Science 294, 858–862 (2001).
Lee, R. C. & Ambros, V. An extensive class of small RNAs in Caenorhabditis elegans. Science 294, 862–864 (2001).
Fire, A. et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391, 806–811 (1998).
Lee, Y. et al. MicroRNA genes are transcribed by RNA polymerase II. EMBO J. 23, 4051–4060 (2004).
Aravin, A. A., Hannon, G. J. & Brennecke, J. The Piwi-piRNA pathway provides an adaptive defense in the transposon arms race. Science 318, 761–764 (2007).
Lee, Y., Jeon, K., Lee, J. T., Kim, S. & Kim, V. N. MicroRNA maturation: stepwise processing and subcellular localization. EMBO J. 21, 4663–4670 (2002).
Lee, Y. et al. The nuclear RNase III Drosha initiates microRNA processing. Nature 425, 415–419 (2003).
Basyuk, E., Suavet, F., Doglio, A., Bordonne, R. & Bertrand, E. Human let-7 stem-loop precursors harbor features of RNase III cleavage products. Nucleic Acids Res. 31, 6593–6597 (2003).
Yi, R., Qin, Y., Macara, I. G. & Cullen, B. R. Exportin-5 mediates the nuclear export of pre-microRNAs and short hairpin RNAs. Genes Dev. 17, 3011–3016 (2003).
Lund, E., Guttinger, S., Calado, A., Dahlberg, J. E. & Kutay, U. Nuclear export of microRNA precursors. Science 303, 95–98 (2004).
Bohnsack, M. T., Czaplinski, K. & Gorlich, D. Exportin 5 is a RanGTP-dependent dsRNA-binding protein that mediates nuclear export of pre-miRNAs. RNA 10, 185–191 (2004).
Ketting, R. F. et al. Dicer functions in RNA interference and in synthesis of small RNA involved in developmental timing in C. elegans. Genes Dev. 15, 2654–2659 (2001).
Grishok, A. et al. Genes and mechanisms related to RNA interference regulate expression of the small temporal RNAs that control C. elegans developmental timing. Cell 106, 23–34 (2001).
Bernstein, E., Caudy, A. A., Hammond, S. M. & Hannon, G. J. Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature 409, 363–366 (2001).
Hutvagner, G. et al. A cellular function for the RNA-interference enzyme Dicer in the maturation of the let-7 small temporal RNA. Science 293, 834–838 (2001).
Hammond, S. M., Bernstein, E., Beach, D. & Hannon, G. J. An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 404, 293–296 (2000).
Zamore, P. D., Tuschl, T., Sharp, P. A. & Bartel, D. P. RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell 101, 25–33 (2000).
Hamilton, A. J. & Baulcombe, D. C. A species of small antisense RNA in posttranscriptional gene silencing in plants. Science 286, 950–952 (1999).
Elbashir, S. M., Lendeckel, W. & Tuschl, T. RNA interference is mediated by 21- and 22-nucleotide RNAs. Genes Dev. 15, 188–200 (2001).
Leuschner, P. J., Ameres, S. L., Kueng, S. & Martinez, J. Cleavage of the siRNA passenger strand during RISC assembly in human cells. EMBO Rep. 7, 314–320 (2006).
Matranga, C., Tomari, Y., Shin, C., Bartel, D. P. & Zamore, P. D. Passenger-strand cleavage facilitates assembly of siRNA into Ago2-containing RNAi enzyme complexes. Cell 123, 607–620 (2005).
Rand, T. A., Petersen, S., Du, F. & Wang, X. Argonaute2 cleaves the anti-guide strand of siRNA during RISC activation. Cell 123, 621–629 (2005).
Hammond, S. M., Boettcher, S., Caudy, A. A., Kobayashi, R. & Hannon, G. J. Argonaute2, a link between genetic and biochemical analyses of RNAi. Science 293, 1146–1150 (2001).
Zeng, Y., Yi, R. & Cullen, B. R. MicroRNAs and small interfering RNAs can inhibit mRNA expression by similar mechanisms. Proc. Natl Acad. Sci. USA 100, 9779–9784 (2003).
Hutvagner, G. & Zamore, P. D. A microRNA in a multiple-turnover RNAi enzyme complex. Science 297, 2056–2060 (2002).
Doench, J. G., Petersen, C. P. & Sharp, P. A. siRNAs can function as miRNAs. Genes Dev. 17, 438–442 (2003).
Song, J. J., Smith, S. K., Hannon, G. J. & Joshua-Tor, L. Crystal structure of Argonaute and its implications for RISC slicer activity. Science 305, 1434–1437 (2004).
Liu, J. et al. Argonaute2 is the catalytic engine of mammalian RNAi. Science 305, 1437–1441 (2004).
Haley, B. & Zamore, P. D. Kinetic analysis of the RNAi enzyme complex. Nat. Struct. Mol. Biol. 11, 599–606 (2004).
Auyeung, V. C., Ulitsky, I., McGeary, S. E. & Bartel, D. P. Beyond secondary structure: primary-sequence determinants license pri-miRNA hairpins for processing. Cell 152, 844–858 (2013).
Fellmann, C. et al. An optimized microRNA backbone for effective single-copy RNAi. Cell Rep. http://dx.doi.org/10.1016/j.celrep.2013.11.020 (2013).
Moffat, J. et al. A lentiviral RNAi library for human and mouse genes applied to an arrayed viral high-content screen. Cell 124, 1283–1298 (2006).
Berns, K. et al. A large-scale RNAi screen in human cells identifies new components of the p53 pathway. Nature 428, 431–437 (2004).
Cheloufi, S., Dos Santos, C. O., Chong, M. M. & Hannon, G. J. A dicer-independent miRNA biogenesis pathway that requires Ago catalysis. Nature 465, 584–589 (2010).
Cifuentes, D. et al. A novel miRNA processing pathway independent of Dicer requires Argonaute2 catalytic activity. Science 328, 1694–1698 (2010).
Yang, J. S. et al. Conserved vertebrate miR-451 provides a platform for Dicer-independent, Ago2-mediated microRNA biogenesis. Proc. Natl Acad. Sci. USA 107, 15163–15168 (2010).
Kumar, M. S. et al. Dicer1 functions as a haploinsufficient tumor suppressor. Genes Dev. 23, 2700–2704 (2009).
Acknowledgements
We thank G. J. Hannon, S. J. Elledge and J. Zuber for continuous support and valuable discussions on state-of-the-art RNAi. We also thank S. Mayack for critical reading of the manuscript and apologize to authors whose work was not cited owing to space constraints. S.W.L. is an investigator of the Howard Hughes Medical Institute.
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C.F. and S.W.L. are founders of Mirimus Inc., and C.F. is an employee of Mirimus Inc., a company that has licensed some of the shRNA technology discussed in this work.
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Fellmann, C., Lowe, S. Stable RNA interference rules for silencing. Nat Cell Biol 16, 10–18 (2014). https://doi.org/10.1038/ncb2895
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DOI: https://doi.org/10.1038/ncb2895
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