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Status |
Public on May 16, 2014 |
Title |
Control matched sample pair brain dorsolateral prefrontal cortex NY_BA9 #10 |
Sample type |
RNA |
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Source name |
Control sample, brain dorsolateral prefrontal cortex
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Organism |
Homo sapiens |
Characteristics |
tissue: brain dorsolateral prefrontal cortex disease state: Control
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Treatment protocol |
not applicable
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Growth protocol |
not applicable
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was extracted from frozen samples stored in TRIzol (Invitrogen, Carlsbad, Calif.) and processed for microarray analysis according to the microarray manufacturer’s protocol (Affymetrix, Santa Clara, Calif.)
|
Label |
biotin
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Label protocol |
According to standard affymetrix protocol: • 10ul Purified cDNA10ul • DEPC H2O • 4ul 10x IVT Labeling Buffer • 12ul IVT Labeling NTP mix • 4ul IVT Labeling Enzyme mix - Incubate at 37C 16 hour in an incubating oven or thermal cycler with heated lid to avoid condensation. - Purify labeled cRNA generated from either IVT labeling reaction using RNeasy RNA Purification Mini kit (Qiagen) using the RNA Cleanup Protocol according to manufacturers instructions. - Quantitate Biotin labeled cRNA using 260nm/280nm spectrophotometric assay.
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Hybridization protocol |
Samples were hybridized using Affymetrix hybridization kit materials: • Heat the hybridization cocktail to 99°C for 5 minutes in a heat block • Incubate the probe array filled with Pre-Hybridization Mix at 45°C for 10 minutes with rotation • Transfer the hybridization cocktail that has been heated at 99°C, to a 45°C heat block for 5 minutes • Spin the hybridization cocktail at maximum speed in a microcentrifuge for 5 minutes to collect any insoluble material from the hybridization mixture • Remove the array from the hybridization oven. Vent the array with a clean pipette tip and extract the Pre-Hybridization Mix from the array with a micropipettor. Refill the array with the appropriate volume of the clarified hybridization cocktail, avoiding any insoluble matter at the bottom of the tube • Place probe array into the hybridization oven, set to 45°C • To avoid stress to the motor, load probe arrays in a balanced configuration around the axis. Rotate at 60 rpm • Hybridize for 16 hours • During the latter part of the 16-hour hybridization, proceed to the GeneChip® Expression Wash, Stain and Scan User Manual, P/N 702731, to prepare reagents for the washing and staining steps required immediately after completion of hybridization
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Scan protocol |
Affymetrix GeneChIP Scanner 3000 7G
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Description |
dissected from frozen coronal blocks ~2–3 cm caudal to the temporal pole
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Data processing |
Probeset signals (i.e., transcript levels) were extracted with the Affymetrix GCOS software. For statistical analysis, log2- transformed probeset signal intensities were extracted and normalized with the robust multiarray average (GC-RMA) algorithm
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Submission date |
Jan 30, 2014 |
Last update date |
May 16, 2014 |
Contact name |
Etienne Sibille |
E-mail(s) |
etienne.sibille@camh.ca
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Phone |
412-624-0804
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Organization name |
University of Pittsburgh
|
Department |
Psychiatry
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Street address |
450 Technology Drive, Suite 231
|
City |
Pittsburgh |
State/province |
PA |
ZIP/Postal code |
15219 |
Country |
USA |
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Platform ID |
GPL96 |
Series (1) |
GSE54570 |
Expression data from human brain dorsolateral prefrontal cortex - including control samples and samples with major depression disorders (26 samples NY_BA9) |
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