The fractional isotopomer distribution according to GC-MS was corrected for fractional distribution

hput approach such as secondgeneration sequencing technology. Delineating the overall gene expression profile in the brain of the HIV-1Tg rat will help to identify the mechanisms involved in HIV-1 neuropathology and allow for the development of efficient therapy for cognitive deficits and other neuropsychiatric disorders associated with HIV-1 infection. primers were ligated to the ends of the DNA fragments. The ligated products were purified on 2% agarose gels, and 200250bp fragments were selected for downstream enrichment by 15 cycles of PCR followed by purification using a QIAquick PCR purification kit. The enriched libraries were diluted with elution buffer to a final concentration of 10 nM. Each sample was subjected to 50 cycles of sequencing from both ends in one lane of an Illmina Hiseq2000 Sequencer. Pre-processing and Mapping of RNA-seq Reads using TopHat The extraction of 50-bp length paired-end reads was achieved using CASAVA. For each sample, reads with a quality score of $Q30 that passed filtering were used to generate a complete FASTQ file, which was then mapped to UCSC Rat reference using TopHat with the default parameter setting of 40 alignments per read and up to 2 mismatches per alignment. The sequence alignment files were analyzed using RSeQC package for quality control analysis, which includes the mRNA fragment insert size, base quality distribution, reads mapping distribution, and splicing distribution analysis. The resulting aligned reads were then analyzed with Cufflinks suite , which assembles the aligned reads into transcripts and measures their relative abundance. The expression of each transcript was quantified as the number of reads mapping to a gene divided by the gene length in kilobases and the total number of mapped reads in millions, which is called fragments per kilobase of exon per million fragments mapped. All the junctions identified by Cufflink were compared on the basis of the junction and splicing site provided by reference transcript annotation GTF files to identify known and novel junctions. Then, Cuffcompare merged all the transcripts from different samples to a final transcript annotation GTF file, reported changes in the relative abundance of transcripts sharing a common transcription start site, and indicated the relative abundance of the primary transcripts of each gene crossing all the samples. Materials and Methods Animals The Animal Care and Use Committee of both the Seton Hall University and University of Virginia approved this study. Adult male HIV-1Tg rats and F344 background control rats were purchased from Harlan Inc.. All rats were double housed in standard plastic cages and maintained in a temperature-controlled environment with a 12 h light/dark cycle and fed a standard rat diet and water ad libitum. The animals were monitored daily, and their 16483784 cage bedding was changed twice a week. All animals were participants in a previously reported behavioral study. All experimental 18772318 procedures were conducted during the light cycle in accordance with the Animal Care and Use Committees of both participating institutions. Tissue Collection Using a rat brain order Go 6983 matrix, slices of approximately 1 mm were taken from each brain, and the slices that contained the PFC, HIP, and dorsal STR were identified according to a rat brain atlas. Tissues from specific regions of interest were collected bilaterally from each brain using a 3.00-mm Harris Micro-Punch and stored at 280uC until use. Gene Annotation and Ex

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