Qiaamp dna mini protocol

Details have been reported previously [14 ]. Briefly, 100 mg sections of frozen dorsolateral prefrontal cortex were obtained from autopsied subjects. Sections were thawed on ice, and the gray matter carefully dissected from the white matter. We extracted DNA with the Qiagen (cat: 51306) QIAamp DNA mini protocol. We used 16uL of DNA at a concentration of 50ng/uL as measured by PicoGreen on the Broad Institute's Genomics Platform for the Illumina InfiniumHumanMethylation450 bead chip assay. The DNA Methylation level for each CpG site is presented as a Beta value (β), which is calculated as the ratio of signal from the methylated probe to the sum of both methylated and unmethylated probes and ranges from 0 to 1. Missing values were imputed using a K-nearest neighboring algorithm with k=100.

Details on RNAseq and methylation data are published (De Jager et al., 2014 (link); Ng et al., 2017 ). Briefly, RNA from 168 individuals was extracted from DLPFC with the miRNeasy mini kit (Qiagen, Venlo, Netherlands) and the RNase free DNase Set (Qiagen, Vento, Netherlands). RNA concentration was quantified using Nanodrop (Thermo Fisher Scientific, Waltham, MA), and RNA quality was assessed using an Agilent Bioanalyzer. RNAseq was performed using Illumina HiSeq with 101 bp paired-end reads with an average depth of 90m reads. The trimmed reads were aligned to the reference genome using Bowtie and the expression FPKM values were estimated using RSEM; see supplement for normalization details.
DNA from 222 individuals was extracted from DLPFC using the Qiagen QIAamp DNA mini protocol. DNA methylation data were generated using Illumina Infinium HumanMethylation450k Bead Chip assay. Raw data were further processed using Methylation Module v1.8 from the Illumina Genome Studio software suite to generate a beta value for each cytosine guanine dinucleotide (CpG), see supplement for normalization details.

100 mg sections of frozen dorsolateral prefrontal cortex were obtained from each of 761 deceased subjects from the ROS and MAP studies based at the Rush Alzheimer's Disease Center. These sections were thawed on ice, and the gray matter was carefully dissected from the white matter. DNA extraction was performed using the Qiagen (cat: 51306) QIAamp DNA mini protocol. The Qubit 2.0 Fluorometer was used to quantitate the DNA. 16uL of DNA at a concentration of 50ng/uL as measured by PicoGreen, was used by the Broad Institute's Genomics Platform for data generation by the Illumina InfiniumHumanMethylation450 bead chip assay. The platform produces a data file by implementing the recommended procedures of the proprietary Illumina GenomeStudio software, which includes color channel normalization and background removal. All data generation was conducted by laboratory personnel who were blinded as to the clinical and neuropathological phenotypes of each subject