age: 92 days sex: Female tissue: dorsolateral prefrontal cortex of the brain
Biomaterial provider
NICHDBB-Baltimore
Treatment protocol
All human postmortem brain tissue samples were obtained from the NICHD Brain and Tissue Bank for Developmental Disorders (NICHDBB)(Baltimore, MD, USA). All subjects were defined as normal controls by forensic pathologists at the NICHDBB. No subjects with prolonged agonal state were used. Chimpanzee samples were obtained from the Yerkes Primate Center (Atlanta, GA, USA), from the Biomedical Primate Research Centre (Rijswijk, Netherlands) and from the Anthropological Institute of the University of Zurich (Switzerland). The majority of chimpanzees used in this study belonged to the Western chimpanzee population. Rhesus macaque samples were obtained from the German Primate Center (DPZ)(Goettingen, Germany). All non-human primates used in this study suffered sudden deaths for reasons other than their participation in this study and without any relation to the tissue used. The human prefrontal cortex samples were taken from the middle third of the middle frontal gyrus: a cortical region approximately corresponding to Brodmann area 46, and from the equivalent region in the non-human primates. For all samples special care was taken to dissect primarily grey matter (in the order of 90-95% of the total sample). The human caudate nucleus samples were taken from the central part of the nucleus in order to avoid any cross-contamination by adjacent brain structures. See publication for a more detailed description of the samples.
Extracted molecule
total RNA
Extraction protocol
Trizol extraction of total RNA from 100 mg of tissue was performed according to the manufacturer's instructions.
Label
biotin
Label protocol
Biotinylated cRNA were prepared from 1 microg. total RNA following standard Affymetrix protocols.
Hybridization protocol
Hybridization to Affymetrix HG-U133 Plus 2.0 GeneChip arrays was carried out following standard Affymetrix protocols.
Scan protocol
GeneChips were scanned using the Hewlett-Packard GeneChip Scanner 3000.
Description
Gene expression data from post-mortem dorsolateral prefrontal cortex of a 0.3 years old human individual
Data processing
Affymetrix microarray image data were collected with Affymetrix GeneChip Operating Software version 1.1 using default parameters. For summarizing the expression data, we used custom chip definition files (Custom CDF, v. 11) based on Ensembl gene annotation rather than Affymetrix annotation (Dai et al., PMID: 16284200). Further, we only accepted probes that perfectly matched both the human, the chimpanzee and the rhesus macaque genomes at a single location (hg18, panTro2, and rheMac2, respectively; see Khaitovich et al., PMID: 16141373), which correspond to 31% of probes on the microarray. Further, we did not include probesets with less than 8 probes in our analysis. The list of exluded probes is supplied as supplement to the CDF file. This list can be used in combination with the removeprobe function in the customCDF package (http://arrayanalysis.mbni.med.umich.edu/MBNIUM.html#CustomCDF) to create CDF environments with masked probes. We used the R Bioconductor 'affy' software package for further data analysis. Signal intensities were calculated with the 'rma' function using default parameters, including quantile normalization and log2 transformation. Detection p-values were calculated with the 'mas5calls' function using default parameters. Note that, for analyses of gene expression changes during postnatal development in humans and chimpanzees only, it is advisable to use CDF files masked for mismatches to the human and chimpanzee genomes, and for analyses involving humans only, to use non-masked CDF files. This is because masking out probes strongly decreases statistical power.
