Project description:Chromatin immunoprecipitation analysis of CENH3 in the Arabidopsis thaliana accessions Col-0, Ler-0, Cvi-0 and Tanz-1 was performed in order to align reads to PacBio HiFi genome assemblies which contain complete centromere repeat arrays.
Project description:PacBio HiFi sequencing was performed on 48 barcoded patients' genomic DNA after a telobait-capture protocol to enrich for telomeric regions. The sequencing reads of each patient were de-multiplexed and presented as patient-specific PacBio CCS BAM files.
Project description:Different Library Sample Preparation (LSP) allow the detection of a large common set of isoforms. However, each LSP also detects a smaller set of isoforms which are characterized both by lower coverage and lower FPKM than that observed for the common ones among LSPs. This characteristic is particularly critical in case of low input RNA NuGEN v2 LSP. The effect of statistical detection of alternative splicing considering low input LSP (NuGEN v2) with respect to high input LSP (TruSeq) was studied using a benchmark dataset, in which both synthetic reads and reads generated from high and low input LSPs were spiked-in. Statistical detection of alternative splicing was done using prototypes of bioinformatics analysis for isoform-reconstruction and exon-level analysis. Each available sample contains a total of 5 paired end replicates. 3 samples contain increasing numbers of spiked-in reads (20, 40, 80 millions) from NuGENv2 library preparation kit on a common TruSeq 1000ng background. 3 additional samples were built with the same approach, but spiked-in reads were collected from a TruSeq-based experiment. The remaining 6 samples follow the same approach of the previous 6, but the common background is based on a TruSeq library preparation on 100ng of material
Project description:We used PacBio data to identify more reliable transcripts from hESC, based on which we can estimate gene/transcript abundance better from Illumina data. PacBio long reads and Illumina short reads were generated from the same hESC cell line H1. PacBio reads were error-corrected by Illumina reads to identify transcripts. rSeq is used to estimate gene/transcript abundance of the identified transcriptome.
Project description:Different Library Sample Preparation (LSP) allow the detection of a large common set of isoforms. However, each LSP also detects a smaller set of isoforms which are characterized both by lower coverage and lower FPKM than that observed for the common ones among LSPs. This characteristic is particularly critical in case of low input RNA NuGEN v2 LSP. The effect of statistical detection of alternative splicing considering low input LSP (NuGEN v2) with respect to high input LSP (TruSeq) was studied using a benchmark dataset, in which both synthetic reads and reads generated from high and low input LSPs were spiked-in. Statistical detection of alternative splicing was done using prototypes of bioinformatics analysis for isoform-reconstruction and exon-level analysis.
Project description:Advances in Next Generation Sequencing (NGS) have made available a wealth of information that had previously been inaccessible to researchers and clinicians. NGS has been applied to understand genomic, transcriptomic, and epigenomic changes and gained traction as a significant tool capable of accelerating diagnosis, prognosis, and biomarker discovery. However, these NGS assays have yet to be practical methods for patient stratification or diagnosis because of the gap between the tiny quantities of biomaterials provided by a clinical sample and the large DNA input required by most of these assays. Current library preparation methodologies typically require large input amounts of DNA and a long and complicated manual process. Here we present a microfluidic reactor system for NGS library preparation, capable of reducing the number of pipetting steps significantly, automating much of the process, while supporting extremely low DNA input requirement (10 pg per library). This largely automated technology will allow for low-input preparations of 8 libraries simultaneously while reducing batch to batch variation and operator hands-on time.