Mammalian RNAi: a practical guide.

Silencing of gene expression by RNA interference (RNAi) has become a powerful tool for the functional annotation of the Caenorhabditis elegans and Drosophila melanogaster genomes. Recent advances in the design and delivery of targeting molecules now permit efficient and highly specific gene silencing in mammalian systems as well. RNAi offers a simple, fast, and cost-effective alternative to existing gene targeting technologies both in cell-based and in vivo settings. Synthetic small interfering RNA (siRNA) and retroviral short hairpin RNA (shRNA) libraries targeting thousands of human and mouse genes are publicly available for high-throughput genetic screens, and knockdown animals can be rapidly generated by lentivirus-mediated transgenesis. RNAi also holds great promise as a novel therapeutic approach. This review provides insight into the current gene silencing techniques in mammalian systems.

[1]  G. Hannon,et al.  Unlocking the potential of the human genome with RNA interference , 2004, Nature.

[2]  Patrick J. Paddison,et al.  An epi-allelic series of p53 hypomorphs created by stable RNAi produces distinct tumor phenotypes in vivo , 2003, Nature Genetics.

[3]  Anton P. McCaffrey,et al.  Inhibition of hepatitis B virus in mice by RNA interference , 2003, Nature Biotechnology.

[4]  David L. Lewis,et al.  Efficient delivery of siRNA for inhibition of gene expression in postnatal mice , 2002, Nature Genetics.

[5]  T. Rana,et al.  siRNA function in RNAi: a chemical modification analysis. , 2003, RNA.

[6]  Wei Ge,et al.  Synthetic shRNAs as potent RNAi triggers , 2005, Nature Biotechnology.

[7]  P. Sandy,et al.  Rnai as an experimental and therapeutic tool to study and regulate physiological and disease processes. , 2005, Annual review of physiology.

[8]  T. Du,et al.  Asymmetry in the Assembly of the RNAi Enzyme Complex , 2003, Cell.

[9]  S. Ohashi,et al.  Sequence‐specific gene silencing in murine muscle induced by electroporation‐mediated transfer of short interfering RNA , 2004, The journal of gene medicine.

[10]  Matthias John,et al.  Therapeutic silencing of an endogenous gene by systemic administration of modified siRNAs , 2004, Nature.

[11]  M. Wiznerowicz,et al.  Conditional Suppression of Cellular Genes: Lentivirus Vector-Mediated Drug-Inducible RNA Interference , 2003, Journal of Virology.

[12]  Nili Jin,et al.  Gene silencing in mammalian cells by PCR‐based short hairpin RNA , 2003, FEBS letters.

[13]  Judy Lieberman,et al.  RNA interference targeting Fas protects mice from fulminant hepatitis , 2003, Nature Medicine.

[14]  Patrick J. Paddison,et al.  A resource for large-scale RNA-interference-based screens in mammals , 2004, Nature.

[15]  Patrick J. Paddison,et al.  RNA interference: the new somatic cell genetics? , 2002, Cancer cell.

[16]  T. Tuschl,et al.  Analysis of gene function in somatic mammalian cells using small interfering RNAs. , 2002, Methods.

[17]  René Bernards,et al.  New tools for functional mammalian cancer genetics , 2003, Nature Reviews Cancer.

[18]  Robert M Sears,et al.  Local gene knockdown in the brain using viral-mediated RNA interference , 2003, Nature Medicine.

[19]  F. Leenders,et al.  Inducible shRNA expression for application in a prostate cancer mouse model. , 2003, Nucleic acids research.

[20]  A. Fire,et al.  Specific inhibition of gene expression by small double-stranded RNAs in invertebrate and vertebrate systems , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[21]  Ye Ding,et al.  Sfold web server for statistical folding and rational design of nucleic acids , 2004, Nucleic Acids Res..

[22]  G. Hannon,et al.  Inducible, reversible, and stable RNA interference in mammalian cells , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[23]  T. Pawson,et al.  Transgenic RNA interference in ES cell–derived embryos recapitulates a genetic null phenotype , 2003, Nature Biotechnology.

[24]  J. Rossi,et al.  Functional siRNA expression from transfected PCR products. , 2002, RNA.

[25]  S. Ishii,et al.  Generation of Ski-knockdown mice by expressing a long double-strand RNA from an RNA polymerase II promoter. , 2003, Genes & development.

[26]  H. Himmelbauer,et al.  An endoribonuclease-prepared siRNA screen in human cells identifies genes essential for cell division , 2004, Nature.

[27]  A. Harel-Bellan,et al.  Conditional gene knock‐down by CRE‐dependent short interfering RNAs , 2004, EMBO reports.

[28]  C. Cepko,et al.  Electroporation and RNA interference in the rodent retina in vivo and in vitro , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[29]  B. Li,et al.  Expression profiling reveals off-target gene regulation by RNAi , 2003, Nature Biotechnology.

[30]  X. Coumoul,et al.  Inducible suppression of Fgfr2 and Survivin in ES cells using a combination of the RNA interference (RNAi) and the Cre-LoxP system. , 2004, Nucleic acids research.

[31]  Vivek Mittal,et al.  Improving the efficiency of RNA interference in mammals , 2004, Nature Reviews Genetics.

[32]  T. Tuschl,et al.  Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells , 2001, Nature.

[33]  Michael T. McManus,et al.  A lentivirus-based system to functionally silence genes in primary mammalian cells, stem cells and transgenic mice by RNA interference , 2003, Nature Genetics.

[34]  R. Schultz,et al.  Transgenic RNAi in mouse oocytes: a simple and fast approach to study gene function. , 2003, Developmental biology.

[35]  M. Amarzguioui,et al.  Tolerance for mutations and chemical modifications in a siRNA. , 2003, Nucleic acids research.

[36]  J. Nunemacher,et al.  Optimal management of giant cell arteritis and polymyalgia rheumatica , 2012, Therapeutics and clinical risk management.

[37]  A. Klippel,et al.  Structural variations and stabilising modifications of synthetic siRNAs in mammalian cells. , 2003, Nucleic acids research.

[38]  M. Okabe,et al.  Small interfering RNA and gene silencing in transgenic mice and rats , 2002, FEBS letters.

[39]  H. Soifer,et al.  siRNA target site secondary structure predictions using local stable substructures , 2005, Nucleic acids research.

[40]  S. Jayasena,et al.  Functional siRNAs and miRNAs Exhibit Strand Bias , 2003, Cell.

[41]  P. Sharp,et al.  Cre-lox-regulated conditional RNA interference from transgenes. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[42]  Serge Batalov,et al.  Identification of modulators of TRAIL-induced apoptosis via RNAi-based phenotypic screening. , 2003, Molecular cell.

[43]  D. Dykxhoorn,et al.  Killing the messenger: short RNAs that silence gene expression , 2003, Nature Reviews Molecular Cell Biology.

[44]  F. Holstege,et al.  Specific inhibition of gene expression using a stably integrated, inducible small‐interfering‐RNA vector , 2003, EMBO reports.

[45]  Patrick J. Paddison,et al.  Production of complex nucleic acid libraries using highly parallel in situ oligonucleotide synthesis , 2004, Nature Methods.

[46]  H. Higashi,et al.  Conditional gene silencing utilizing the lac repressor reveals a role of SHP‐2 in cagA‐positive Helicobacter pylori pathogenicity , 2004, Cancer science.

[47]  M. Manoharan,et al.  RNA interference in vivo: toward synthetic small inhibitory RNA-based therapeutics. , 2005, Methods in enzymology.

[48]  Peter A. Jones,et al.  Establishment of conditional vectors for hairpin siRNA knockdowns. , 2003, Nucleic acids research.

[49]  G. Stamatoyannopoulos,et al.  Down-regulation of CXCR4 by inducible small interfering RNA inhibits breast cancer cell invasion in vitro. , 2003, Cancer research.

[50]  T. Rana,et al.  RNAi in human cells: basic structural and functional features of small interfering RNA. , 2002, Molecular cell.

[51]  Vivek Mittal,et al.  High-throughput selection of effective RNAi probes for gene silencing. , 2003, Genome research.

[52]  Inder M Verma,et al.  A general method for gene knockdown in mice by using lentiviral vectors expressing small interfering RNA , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[53]  Peter G Schultz,et al.  An approach to genomewide screens of expressed small interfering RNAs in mammalian cells , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[54]  Harvey F Lodish,et al.  Small interfering RNA production by enzymatic engineering of DNA (SPEED). , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[55]  A. Reynolds,et al.  Rational siRNA design for RNA interference , 2004, Nature Biotechnology.

[56]  F. Buchholz,et al.  Short RNA duplexes produced by hydrolysis with Escherichia coli RNase III mediate effective RNA interference in mammalian cells , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[57]  Helen M Blau,et al.  Restriction enzyme–generated siRNA (REGS) vectors and libraries , 2004, Nature Genetics.

[58]  C. Auffray,et al.  The I.M.A.G.E. Consortium: an integrated molecular analysis of genomes and their expression. , 1996, Genomics.

[59]  Joshua T. Jones,et al.  Recombinant Dicer efficiently converts large dsRNAs into siRNAs suitable for gene silencing , 2003, Nature Biotechnology.

[60]  M. Iino,et al.  Enzymatic production of RNAi libraries from cDNAs , 2004, Nature Genetics.

[61]  I. Verma,et al.  CRE recombinase-inducible RNA interference mediated by lentiviral vectors. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[62]  Michael T. McManus,et al.  Small Interfering RNA-Mediated Gene Silencing in T Lymphocytes1 , 2002, The Journal of Immunology.

[63]  T. Tuschl,et al.  Mechanisms of gene silencing by double-stranded RNA , 2004, Nature.

[64]  Thomas Tuschl,et al.  siRNAs: applications in functional genomics and potential as therapeutics , 2004, Nature Reviews Drug Discovery.

[65]  Sangdun Choi,et al.  Synthetic dsRNA Dicer substrates enhance RNAi potency and efficacy , 2005, Nature Biotechnology.

[66]  Haibin Xia,et al.  siRNA-mediated gene silencing in vitro and in vivo , 2002, Nature Biotechnology.

[67]  K. Taira,et al.  Control of siRNA expression using the Cre-loxP recombination system. , 2004, Nucleic acids research.

[68]  David R Corey,et al.  RNA interference in mammalian cells by chemically-modified RNA. , 2003, Biochemistry.

[69]  Reuven Agami,et al.  A large-scale RNAi screen in human cells identifies new components of the p53 pathway , 2004, Nature.