title: Exposure to dsRNA elicits RNA interference in Brachionus manjavacas (Rotifera).
abstract: RNA interference (RNAi) is a powerful technique for functional genomics, yet no studies have reported its successful application to zooplankton. Many zooplankton, particularly microscopic metazoans of phylum Rotifera, have unique life history traits for which genetic investigation has been limited. In this paper, we report the development of RNAi methods for rotifers, with the exogenous introduction of double-stranded RNA (dsRNA) through the use of a lipofection reagent. Transfection with dsRNA for heat shock protein 90, the membrane-associated progesterone receptor, and mitogen-activated protein kinase significantly increased the proportion of non-reproductive females. Additionally, a fluorescence-based lectin binding assay confirmed the significant suppression of four of six glycosylation enzymes that were targeted with dsRNA. Suppression of mRNA transcripts was confirmed with quantitative PCR. Development of RNAi for rotifers promises to enhance the ability for assessing genetic regulation of features critical to their life history and represents a key step toward functional genomics research in zooplankton.
authors.affiliation: School of Biology, Georgia Institute of Technology, 311 Ferst Drive, Atlanta, GA 30332-0230, USA. terry.snell@biology.gatech.edu
authors.forename: Terry W
authors.initials: TW
authors.lastname: Snell
authors.forename: Tonya L
authors.initials: TL
authors.lastname: Shearer
authors.forename: Hilary A
authors.initials: HA
authors.lastname: Smith
mesh_terms: D000818:Animals; D005260:Female; D015870:Gene Expression; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques / Q000379:methods*; D020868:Gene Silencing; D018841:HSP90 Heat-Shock Proteins / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000008:administration & dosage* / Q000235:genetics / Q000378:metabolism; D012098:Reproduction / Q000235:genetics; D012403:Rotifera / Q000235:genetics*; D014162:Transfection
chemical_list: D018841:HSP90 Heat-Shock Proteins; D012330:RNA, Double-Stranded; D020928:Mitogen-Activated Protein Kinases
references.citation: Adv Drug Deliv Rev. 2005 Apr 5;57(5):733-53
references.pmid: 15757758
references.citation: Evolution. 2002 Jul;56(7):1431-44
references.pmid: 12206243
references.citation: Nature. 2000 Nov 16;408(6810):325-30
references.pmid: 11099033
references.citation: Proc Natl Acad Sci U S A. 1987 Nov;84(21):7413-7
references.pmid: 2823261
references.citation: Genes Dev. 2002 Aug 1;16(15):1884-9
references.pmid: 12154120
references.citation: Methods Mol Biol. 2007;405:23-9
references.pmid: 18369814
references.citation: PLoS One. 2007 Aug 01;2(7):e671
references.pmid: 17668053
references.citation: Nature. 2002 Jul 11;418(6894):244-51
references.pmid: 12110901
references.citation: Nature. 2004 Jul 8;430(6996):161-4
references.pmid: 15241403
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Dev Genes Evol. 2008 Aug;218(8):445-51
references.pmid: 18594859
references.citation: PLoS One. 2007 Aug 01;2(8):e694
references.pmid: 17668071
references.citation: Nature. 2008 Apr 10;452(7188):745-9
references.pmid: 18322464
references.citation: Curr Genet. 2006 Aug;50(2):81-99
references.pmid: 16691418
references.citation: Insect Biochem Mol Biol. 2009 Nov;39(11):824-32
references.pmid: 19815067
references.citation: J Cell Sci. 2002 Jun 15;115(Pt 12):2457-9
references.pmid: 12045215
references.citation: Methods. 2004 Jun;33(2):104-12
references.pmid: 15121164
references.citation: PLoS One. 2007 Jun 06;2(6):e509
references.pmid: 17551589
references.citation: Nature. 2003 Jan 16;421(6920):231-7
references.pmid: 12529635
references.citation: Trends Ecol Evol. 2008 Oct;23(10):578-87
references.pmid: 18715673
references.citation: Nature. 1998 Feb 19;391(6669):806-11
references.pmid: 9486653
references.citation: J Cell Sci. 2006 Mar 1;119(Pt 5):846-57
references.pmid: 16478786
references.citation: Proc Biol Sci. 2009 Apr 7;276(1660):1245-54
references.pmid: 19129141
references.citation: Proc Natl Acad Sci U S A. 2003 Feb 18;100(4):1844-8
references.pmid: 12552109
references.citation: Proc Natl Acad Sci U S A. 2000 Jun 6;97(12):6499-503
references.pmid: 10823906
references.citation: Trends Genet. 2008 Jun;24(6):297-305
references.pmid: 18450316
references.citation: BMC Genomics. 2009 Mar 13;10:108
references.pmid: 19284654
references.citation: Dev Genes Evol. 2003 Apr;213(3):138-41
references.pmid: 12690452
references.citation: BMC Biol. 2009 Sep 09;7:60
references.pmid: 19740420
references.citation: Methods. 2004 Jun;33(2):95-103
references.pmid: 15121163
references.citation: Mol Biol Cell. 2003 Jan;14(1):251-61
references.pmid: 12529441