Comment[ArrayExpressAccession]	E-GEOD-59965
MAGE-TAB Version	1.1			
Public Release Date	2014-09-01
Investigation Title	A trans-generational epigenetic process defines piRNA biogenesis in Drosophila virilis			
Comment[Submitted Name]	A trans-generational epigenetic process defines piRNA biogenesis in Drosophila virilis
Experiment Description	Piwi interacting (pi)RNAs repress diverse transposable elements in the germ cells of metazoans and are essential for fertility in both invertebrates and vertebrates. The precursors of piRNAs are transcribed from distinct genomic regions, the so-called piRNA clusters; however, how piRNA clusters are differentiated from the rest of the genome is not known. To address this question, we studied piRNA biogenesis in two Drosophila  virilis strains that show differential ability to generate piRNAs from several genomic regions. We found that active piRNA biogenesis correlates with high levels of histone 3 lysine 9 trimethylation (H3K9me3) over genomic regions that give rise to piRNAs. Furthermore, piRNA biogenesis in the progeny requires the trans-generational inheritance of an epigenetic signal, presumably in form of homologous piRNAs that are generated in the maternal germline and deposited into the oocyte. The inherited piRNAs enhance piRNA biogenesis by installment of H3K9me3 mark on piRNA clusters and by promoting ping-pong processing of homologous transcripts into mature piRNAs.  We submitted the resequencing data together with the functional genomic datasets because it was generated with the sole purpose of supporting those. The SRA accession numbers are SRR1536176 and SRR1536175. ChIP-seq against H3K9me3 and Pol2, Total RNA-seq, in Drosophila virilis Strain9 and Strain160 as well as crosses between them			
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl		
Person Last Name	Mairnov	Marinov	Thomas	Aravin
Person First Name	Georgi	Georgi	Adrien	Alexei
Person Mid Initials	K	K	L	A
Person Email	georgi@caltech.edu			
Person Affiliation	California Institute of Technology			
Person Phone	6262543740			
Person Address	California Institute of Technology, 1200 E California Blvd, Pasadena, California, USA			
Person Roles	submitter			
Protocol Name	P-GSE59965-1	P-GSE59965-2		
Protocol Description	Flies were put on yeast for 2-3 days and 100 ovaries per IP condition were dissected. Ovaries were fixed for 10 min at RT using 1% paraformaldehyde (PFA) followed by 5 min quenching at RT by directly adding glycine (final concentration 25mM) and 3 washes in PBS. Ovaries were slightly dounced in Farnham Buffer (5mM HEPES pH8.0; 85mM KCl; 0.5% NP-40/Igepal; NaCl Protease Inhibitor Cocktail; NaF 10mM; Na3VO4 0.2mM), palleted, followed by strong douncing in RIPA Buffer (20mM Tris pH7.4; 150mM; 1% NP-40/Igepal; 0.5% Sodium Deoxycholate; 0.1% SDS; Protease Inhibitor Cocktail; NaF 10mM; Na3VO4 0.2mM) prior to sonication using a Bioruptor from Diagenode on medium power for 20 cycles (30sec ON, 30sec OFF). Samples were then centrifuged, supernatant collected and pre-cleared for 2h at 4ºC using Dynabeads Protein G (Invitrogen). Antibodies were conjugated to Dynabeads Protein G for 2h at 4ºC. A sample of 5% of the pre-cleared supernatant was saved as input control fraction and the rest was incubated with the antibody-conjugated beads for 2h at 4ºC. The beads were washed 5 times at 4ºC using LiCl IP Buffer (10mM Tris ph7.5; 500mM LiCL; 1% NP-40/Igepal; 1% Sodium Deoxycholate), once with TE. Samples were digested in Proteinase K Buffer (200mM Tris ph7.4; 25mM EDTA; 300mM NaCL; 2% SDS) with 100µg of proteinase K. Samples for 3h at 55ºC and revers-crosslinked overnight at 65ºC. DNA was extracted following standard phenol-chloroform extraction and concentration was measured by Qubit. Libraries were generated following Illumina's DNA Sample Prep Kit protocol	RNA was extracted from 30 flies using standard ribosol RNA extraction. Samples were then rRNA depleted using Ribozero Kit, and DNAse treated prior to library preparation and sequencing.		
Protocol Type	nucleic acid library construction protocol	nucleic acid library construction protocol		
Experimental Factor Name	CHIP-ANTIBODY	STRAIN		
Experimental Factor Type	chip-antibody	strain		
Comment[SecondaryAccession]	GSE59965			
Comment[GEOReleaseDate]	2014-09-01			
Comment[ArrayExpressSubmissionDate]	2014-07-31			
Comment[GEOLastUpdateDate]	2014-09-02			
Comment[AEExperimentType]	RNA-seq of coding RNA			
Comment[AEExperimentType]	ChIP-seq			
Comment[SecondaryAccession]	SRP045147			
SDRF File	E-GEOD-59965.sdrf.txt