title: RNAi-Mediated Inactivation of Autophagy Genes in Caenorhabditis elegans.
abstract: RNA interference (RNAi) is a process that results in the sequence-specific silencing of endogenous mRNA through the introduction of double-stranded RNA (dsRNA). In the nematode Caenorhabditis elegans, RNA inactivation can be used at any specific developmental stage or during adulthood to inhibit a given target gene. Investigators can take advantage of the fact that, in C. elegans, RNAi is unusual in that it is systemic, meaning that dsRNA can spread throughout the animal and can affect virtually all tissues except neurons. Here, we describe a protocol for the most common method to achieve RNAi in C. elegans, which is to feed them bacteria that express dsRNA complementary to a specific target gene. This method has various advantages, including the availability of libraries that essentially cover the whole genome, the ability to treat animals at any developmental stage, and that it is relatively cost effective. We also discuss how RNAi specific to autophagy genes has proven to be an excellent method to study the role of these genes in autophagy, as well as other cellular and developmental processes, while also highlighting the caveats that must be applied.
authors.affiliation: Department of Biology, Queens College-CUNY, Flushing, New York 11367; The Graduate Center, The City University of New York, New York 10016.
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Palmisano
authors.affiliation: Department of Biology, Queens College-CUNY, Flushing, New York 11367; The Graduate Center, The City University of New York, New York 10016.
authors.forename: Alicia
authors.initials: A
authors.lastname: Meléndez
mesh_terms: D000818:Animals; D001343:Autophagy; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000502:physiology*; D055785:Gene Knockdown Techniques / Q000379:methods*; D017238:Genes, Helminth; D034622:RNA Interference
references.citation: Curr Biol. 2002 Aug 6;12(15):1317-9
references.pmid: 12176360
references.citation: Cold Spring Harb Protoc. 2016 Jan 04;2016(1):pdb.prot086504
references.pmid: 26729906
references.citation: Autophagy. 2006 Jul-Sep;2(3):189-99
references.pmid: 16874109
references.citation: Nature. 1998 Feb 19;391(6669):806-11
references.pmid: 9486653
references.citation: Cell. 2001 Nov 16;107(4):465-76
references.pmid: 11719187
references.citation: Gene. 2001 Jan 24;263(1-2):103-12
references.pmid: 11223248
references.citation: Curr Biol. 2011 Sep 27;21(18):1507-14
references.pmid: 21906946
references.citation: Genome Res. 2004 Oct;14(10B):2162-8
references.pmid: 15489339
references.citation: Genome Biol. 2006;7(1):R4
references.pmid: 16507136
references.citation: Cold Spring Harb Protoc. 2016 Jan 04;2016(1):pdb.prot086496
references.pmid: 26729905
references.citation: Nature. 1998 Oct 29;395(6705):854
references.pmid: 9804418
references.citation: PLoS One. 2012;7(5):e35428
references.pmid: 22574120
references.citation: Autophagy. 2011 Apr;7(4):386-400
references.pmid: 21183797
references.citation: Science. 2003 Sep 5;301(5638):1387-91
references.pmid: 12958363
references.citation: Mech Ageing Dev. 1977 Nov-Dec;6(6):413-29
references.pmid: 926867
references.citation: Biol Chem. 2004 Jun;385(6):565-8
references.pmid: 15255192
references.citation: Proc Natl Acad Sci U S A. 2007 Jan 16;104(3):834-9
references.pmid: 17213328
references.citation: Science. 2000 Mar 31;287(5462):2494-7
references.pmid: 10741970
references.citation: Nat Methods. 2010 Jul;7(7):554-9
references.pmid: 20512143
references.citation: Nature. 2004 Feb 12;427(6975):645-9
references.pmid: 14961122
references.citation: Curr Biol. 2000 Feb 24;10(4):169-78
references.pmid: 10704412
references.citation: Genome Biol. 2001;2(1):RESEARCH0002
references.pmid: 11178279
references.citation: Nature. 2005 Jul 28;436(7050):593-7
references.pmid: 16049496
references.citation: Autophagy. 2007 Mar-Apr;3(2):93-5
references.pmid: 17204841
references.citation: PLoS Genet. 2008 Feb;4(2):e24
references.pmid: 18282106
references.citation: Cell. 1999 Oct 15;99(2):123-32
references.pmid: 10535731
references.citation: Gene. 2007 Oct 1;400(1-2):166-73
references.pmid: 17681718
references.citation: Methods Enzymol. 2008;451:493-520
references.pmid: 19185737
references.citation: Science. 2002 Mar 29;295(5564):2456-9
references.pmid: 11834782
references.citation: Nature. 2003 Jan 16;421(6920):231-7
references.pmid: 12529635