Project description:Peltate glandular trichomes from /Mentha spicata /were purified on a Percoll gradient and soluble and membrane proteins were trypsinized and the peptides separated by nano-LC fractionation and analyzed by MALDI-MS/MS. The vast majority of the 1,666 proteins identified was housekeeping proteins or involved in primary metabolism. However, 57 were predicted to be involved in secondary metabolism. Of these, 21 were involved in the synthesis of phenylpropanoids and phenolics and 32 in terpenoid synthesis. Of the 14 membrane transporters identified, the 11 ATP-binding cassette transporters provide good material for assessing whether active transport is required for the transfer of monoterpenoid intermediates between cellular compartments and for the secretion of the final products into the subcuticular storage cavity. In conclusion, this proteome analysis of /M. spicata/ peltate trichomes has identified several candidate proteins that might be involved in terpenoid synthesis and transport. MALDI-MS/MS analyses were performed as described previously (Szopinska et al., 2011). MS data from the soluble and microsomal fractions were pooled and analyzed as a single data set using ProteinPilotTM software (AB SCIEX, v.4.0.8085). Protein identification was based on ParagonTM algorithm v.4.0 and used the Viridiplantae part of the whole NCBInr protein database (downloaded on April 14th, 2013). This algorithm in ProteinPilot was used with “identification” as the sample type, “MMTS” as cysteine modification, “4800 TOF/TOF” as the instrument, and the "Thorough" preset search setting. All reported proteins were identified with 95% or greater confidence, as determined by ProteinPilot unused scores (greater than 1.3) using the most stringent threshold of false positive discovery rate, i.e. below 10%. Protein grouping performed by ProteinPilotTM removed redundant hits.
Project description:Purpose: The goal of this study is to compare endothelial small RNA transcriptome to identify the target of OASL under basal or stimulated conditions by utilizing miRNA-seq. Methods: Endothelial miRNA profilies of siCTL or siOASL transfected HUVECs were generated by illumina sequencing method, in duplicate. After sequencing, the raw sequence reads are filtered based on quality. The adapter sequences are also trimmed off the raw sequence reads. rRNA removed reads are sequentially aligned to reference genome (GRCh38) and miRNA prediction is performed by miRDeep2. Results: We identified known miRNA in species (miRDeep2) in the HUVECs transfected with siCTL or siOASL. The expression profile of mature miRNA is used to analyze differentially expressed miRNA(DE miRNA). Conclusions: Our study represents the first analysis of endothelial miRNA profiles affected by OASL knockdown with biologic replicates.
Project description:A cDNA library was constructed by Novogene (CA, USA) using a Small RNA Sample Pre Kit, and Illumina sequencing was conducted according to company workflow, using 20 million reads. Raw data were filtered for quality as determined by reads with a quality score > 5, reads containing N < 10%, no 5' primer contaminants, and reads with a 3' primer and insert tag. The 3' primer sequence was trimmed and reads with a poly A/T/G/C were removed
Project description:Whole exome sequencing of 5 HCLc tumor-germline pairs. Genomic DNA from HCLc tumor cells and T-cells for germline was used. Whole exome enrichment was performed with either Agilent SureSelect (50Mb, samples S3G/T, S5G/T, S9G/T) or Roche Nimblegen (44.1Mb, samples S4G/T and S6G/T). The resulting exome libraries were sequenced on the Illumina HiSeq platform with paired-end 100bp reads to an average depth of 120-134x. Bam files were generated using NovoalignMPI (v3.0) to align the raw fastq files to the reference genome sequence (hg19) and picard tools (v1.34) to flag duplicate reads (optical or pcr), unmapped reads, reads mapping to more than one location, and reads failing vendor QC.