MAGE-TAB Version	1.1
Investigation Title	TARGET: Kidney - Rhabdoid Tumor (RT) Bisulfite-seq
Experimental Design
Experimental Design Term Source REF
Experimental Factor Name
Experimental Factor Type
Experimental Factor Term Source REF
Person Last Name	NCI Office of Cancer Genomics (OCG)	NCI Center for Biomedical Informatics and Information Technology (CBIIT)	Perlman	Ma	Novik
Person First Name			Elizabeth	Yussanne	Karen
Person Mid Initials				P	L
Person Email	ocg@mail.nih.gov	ncicbiit@mail.nih.gov	eperlman@luriechildrens.org	yma@bcgsc.ca	knovik@bcgsc.ca
Person Phone	+1 301 451 8027	+1 888 478 4423	+1 312 227 3967	+1 604 707 5800 Ext 6082	+1 604 707 8000 Ext 7983
Person Fax	+1 301 480 4368			+1 604 876 3561	+1 604 675 8178
Person Address	31 Center Dr, Rm 10A07, Bethesda, MD 20892	9609 Medical Center Dr, Rockville, MD 20850	225 E Chicago Ave, Chicago, IL 60611	Suite 100-570 West 7th Ave, Vancouver, BC Canada V5Z 4S6	675 West 10th Ave Vancouver, BC Canada V5Z 1L3
Person Affiliation	National Cancer Institute	National Cancer Institute	Ann & Robert H. Lurie Children's Hospital of Chicago	BC Cancer Agency Canada's Michael Smith Genome Sciences Centre	BC Cancer Agency Canada's Michael Smith Genome Sciences Centre
Person Roles	funder;investigator	data coder;curator	investigator	investigator;data analyst;submitter	investigator
Person Roles Term Source REF	EFO;EFO	EFO;EFO	EFO	EFO;EFO;EFO	EFO
Quality Control Type
Quality Control Term Source REF
Replicate Type
Replicate Term Source REF
Normalization Type
Normalization Term Source REF
Date of Experiment
Public Release Date
PubMed ID
Publication DOI
Publication Author List
Publication Title
Publication Status
Publication Status Term Source REF
Experiment Description	"To further explore some more aggressive subtypes in pediatric cancer already under study in OCG, CGCI is partnering with TARGET initiative to support extensive sequencing analysis of three hard to treat childhood cancers: refractory to treatment cases of acute myeloid leukemia and two rare kidney tumors; clear cell sarcoma of the kidney and rhabdoid tumor (RT). Rhabdoid tumors are a rare, very aggressive type of kidney tumor of infancy, and patients typically have a poor prognosis. Up to 40 patient cases of matched tumor and normal samples, as well as some established cell lines, will undergo whole genome sequencing, miRNA-seq and mRNA-seq, DNA methyl-seq and ChIP-seq to generate a comprehensive profile and hopefully facilitate the development of better therapeutic options for this tumor subtype. All cases can be sorted according to data type via the Case Matrix on the TARGET Data Matrix. Please visit the TARGET website listed above for additional information... (for more see dbGaP study page.)"
Protocol Name	nationwidechildrens.org:Protocol:DNA-Extraction-Qiagen-AllPrep:01	bcgsc.ca:Protocol:Bisulfiteseq-LibraryPrep-Illumina:01	bcgsc.ca:Protocol:Bisulfiteseq-Sequence-Illumina-HiSeq2000:01	bcgsc.ca:Protocol:Bisulfiteseq-Sequence-Illumina-HiSeq2500:01	bcgsc.ca:Protocol:Bisulfiteseq-BaseCall-Illumina:01	bcgsc.ca:Protocol:Bisulfiteseq-ReadAlign:01	bcgsc.ca:Protocol:Bisulfiteseq-Methylation:01
Protocol Type	nucleic acid extraction protocol	nucleic acid library construction protocol	nucleic acid sequencing protocol	nucleic acid sequencing protocol	data transformation protocol	data transformation protocol	data transformation protocol
Protocol Term Source REF	EFO	EFO	EFO	EFO	EFO	EFO	EFO
Protocol Description	"Genomic DNA was prepared from using the Qiagen AllPrep DNA/RNA Mini Kit (Qiagen, Valencia, CA, USA). Please see https://ocg.cancer.gov/programs/target/target-methods for full extraction protocol details."	"To track the efficiency of bisulfite conversion, 1ng lambda DNA (Promega) was spiked into 1µg genomic DNA quantified using Qubit fluorometry and arrayed in a 96-well microtitre plate. DNA was sheared to a target size of 300bp using Covaris sonication and the fragments end repaired using DNA ligase and dNTPs at 30oC for 30min. Repaired DNA was purified using a 2:1 AMPure XP beads to sample ratio and eluted in 40µL elution buffer in preparation for A-tailing; the addition of adenosine to the 3’ end of DNA fragments using Klenow fragment and dATP incubated at 37oC for 30min. Following reaction clean-up with magnetic beads, cytosine methylated paired-end adapters (5’-AmCAmCTmCTTTmCmCmCTAmCAmCGAmCGmCTmCTTmCmCGATmCT-3’ and 3’-GAGmCmCGTAAGGAmCGAmCTTGGmCGAGAAGGmCTAG-5’) were ligated to the DNA at 30oC, 20min and adapter flanked DNA fragments bead purified. Prior to bisulfite conversion an aliquot of library fragments was amplified with 10 cycles of PCR and sized on an Agilent Bioanalyzer High Sensitivity DNA chip. Amplicons were observed between 200-700bp in length. Bisulfite conversion of the methylated adapter-ligated DNA fragments was achieved using the EZ Methylation-Gold kit (Zymo Research) following the manufacturer’s protocol. Five cycles of PCR using HiFi polymerase (Kapa Biosystems) was used to enrich the bisulfite converted DNA and introduce fault tolerant hexamer barcode sequences. Post-PCR purification and size-selection of bisulfite converted DNA was performed from pre-cast 8% TBE gels (Invitrogen), extracting the 350-500bp fraction, or 275-425bp fraction if the former was of weak intensity. Gel slurries were added to Spin-X filter tubes (Fisher) and the eluate ethanol precipitated and resuspended in EB. To determine final library concentrations fragment sizes were assessed using a high sensitivity DNA assay (Agilent) and DNA quantified by Qubit fluorometry. Where necessary, libraries were diluted in elution buffer supplemented with 0.1% Tween-20 to achieve a concentration of 8nM for Illumina HiSeq2000/2500 flowcell cluster generation."					
Protocol Parameters					Software Versions		
Protocol Hardware			Illumina HiSeq 2000	Illumina HiSeq 2500			
Protocol Software					Illumina RTA		
Protocol Contact
SDRF File	TARGET_RT_Bisulfite-seq_20190320.sdrf.txt
Term Source Name	NCBITaxon	NCIt	MO	EFO	OBI
Term Source File	http://www.ncbi.nlm.nih.gov/taxonomy	http://ncit.nci.nih.gov/	http://mged.sourceforge.net/ontologies/MGEDontology.php	http://www.ebi.ac.uk/efo	http://purl.obolibrary.org/obo/obi
Term Source Version
Comment[SRA_STUDY]	SRP012005
Comment[BioProject]	PRJNA89539
Comment[dbGaP Study]	phs000470
