Cells grown in RPMI 1640 with 5% FBS (Bio Whittaker, not heat inactivated) and 1% L-glutamine. Cells not grown past 80% confluencey for attached cells, or 0.5 x 10^6 cells for suspended. Trypsinize (for attached cells) cells with 5 ml trypsin-EDTA per T162, for 15 min., at 37C. Spin down suspended cells, and resuspend in ~3ml growth media. Count cells. Pass 1x10^6 cells into new T162's w 30 ml media. Repeat growth cycle until desired amount of cells are available. Cells are not grown past passage 20.
Extracted molecule
genomic DNA
Extraction protocol
Genomic DNA was purified from cells using either the QIAamp DNA Blood Maxi Kit, or the Blood & Cell Culture DNA Maxi Kit (Qiagen Inc., Valencia, CA) according to manufacturer's instructions. Quality was assessed by optical density 260/280 ratio using a spectrophotometer (Beckman-Coulter, Fullerton, CA) and by 0.8% agarose (SeaKem GTG, FMC BioProducts, Rockland, ME) gel electrophoresis in 1x TAE (Roche, Indianapolis, IN).
Label
Biotin
Label protocol
As per manufacturer (Affymetrix)
Hybridization protocol
DNA was restriction digested, PCR amplified, fragmented, labeled and hybridized to each array according to the manufacturer's instructions.
Scan protocol
The Arrays were then washed using Affymetrix fluidics stations, and scanned using the Gene Chip Scanner 3000 7G with AutoLoader.
Description
Hybridized to 250K_Nsp RCO.1-250knsp
Data processing
Genotype call was made by Affymetrix Power Tool (Linux, version 1.12.0) BRLMM algorithm. ROH was inferred by dChip (Version 2010/01) basic-HMM module, and PLINK (Version 1.07) ROH module. dChip ROH Call: L, R, and N; PLINK ROH Call: 0, 1 [dChip.txt, PLINK_coverted.txt, and PLINK_rawData.txt are provided as Series supplementary files.]
