The BAZ1B gene was knocked out by targeting both copies of exon-7 using zinc finger nucleases (D5, F3 and M1 - all independent clones). Clone A6 is targeted at one allele only and is a heterozygous mutant clone.
Growth protocol
Cells were grown in DMEM/F12, supplemented with 10% FBS, Pen/Strep, 2mM L-Glutamine, 0.348% NaBicarbonate. Cells were maintained at 37C in 5% CO2, humidified.
Extracted molecule
total RNA
Extraction protocol
Total RNA was isolated using the Qiagen RNeasy Spin Kit. RNA was treated with DNaseI, before cleaning on an RNeasy spin column. Confirmation of DNA absence was performed by PCR to the repeat element DXZ4, and RNA quality was determined by agarose gel electrophoresis.
Label
Cy3
Label protocol
Labeling was performed at a NimbleGen Systems certified core facility at FSU, FL, USA, following the NimbleGen standard operating protocol. See www.nimblegen.com.
Hybridization protocol
Hybridization was performed at a NimbleGen Systems certified core facility at FSU, FL, USA, following the NimbleGen standard operating protocol. See www.nimblegen.com.
Scan protocol
Scanning was performed at a NimbleGen Systems certified core facilty at FSU, FL, USA, following the NimbleGen standard operating protocol. See www.nimblegen.com.
Description
Parental RPE1 cells.
Data processing
The raw data (.pair file) was subjected to RMA (Robust Multi-Array Analysis; Irizarry et al. Biostatistics 4(2):249), quantile normalization (Bolstad et al. Bioinformatics 19(2):185), and background correction as implemented in the NimbleScan software package, version 2.4.27 (Roche NimbleGen, Inc.).
