• 1 Project Title
• 2 Initial Meetings and Timeline
• 3 Project Description
• 3.1 Goals of the project
• 4 Analysis plan
• 5 Received Data
• 5.1 Sample Summary
• 6 Results Summary
• 6.1 QC steps
• 6.2 Statistics based on per sample analysis
• 7 Appendix

NGS Bioinformatics for mrnaseq sequencing

1. Background

item	description
Disease Type	TEST
Number of Samples	1
Read Length	50
PairedEnd(PE)/SingleRead(SR)	PE
Genome Build (hg18/hg19)	hg19
StartDate	2/25/2014
EndDate	2/25/2014

Note: Further raw NGS data will be used for statistical analysis
Further analysis space:: \\rcfcluster-cifs\NA

Data is available for 60 Days from the Delivered Date at
Delivery space:: \\rcfcluster-cifs\data2\external_data\Bockol_Matthew_m084061\maprseq\delivery

2. Study design

• What are the samples?
• MAPRSeq sample run
• Goals of the project
• Identify how well mrnaseq sequencing worked on these samples,and obtain Gene and exon expression

• Run Name
• test_sample
• Sample Summary



Lane	Index	Sample Names
-	-	SAMPLE1

• QC steps - FastQC-report
• FastQC aims to provide a simple way to do some quality control checks on raw sequence data coming from high throughput sequencing pipelines. It provides a modular set of analyses which you can use to give a quick impression of whether your data has any problems of which you should be aware before doing any further analysis.
  FastQC Reports
• Statistics based on per Sample Analysis (ColumnDescription)
• Analysis is carried out using fastq sequence files as input and generates output tables. For paired-end runs, the tables contain counts for each sample combined from both reads.

sample(s)	total reads	used reads	mapped reads	mapped reads (genome)	mapped reads (junction)	gene count	exon count
SAMPLE1	500,000	499,926	474,913 (95.0)	404,293 (80.9)	70,620 (14.1)	337,660 (67.5)	429,192

• The following three sets of tables are delivered and column description is available in Appendix.
• Exon table: contains counts for the number of times an exon has been detected
• count (raw) = sum of exon read counts
• count (RPKM)
• Gene table: contains counts for the number of times a gene copy has been detected
• count (raw) = sum of exon read counts, with an exception that if reads start in different exons of the same gene twice, these are counted only once for the gene
• Single Nucleotide Variants: contains single nucleotide variants (SNVs) identified per sample
• filter.vcf = lists SNVs identified along with information on whether the variant passed recommended filtering criteria (can be observed at the FILTER field)
• RSeQC Plots:



• Sample Name	Junction Saturation	Splicing Junction	Splicing Event	Inner distance	Read duplication	Gene body coverage
  SAMPLE1



• Tophat fusion (circos plot and raw files):
• Tab delimited fusion results are available here
• Detail fusion report in HTML format is available here

• Circos plot

• Statistics based on per Sample Analysis are recorded in the tab delimited file Details


• IGV Visualization
• The SNV annotation reports (both standard and filtered) include visualization links to IGV to enable a realistic view of the variants. Please follow steps in the following link to setup IGV (takes less than 5 minutes) and utilize this feature. IGV setup for variant visualization

• Tophat 2
• Tophat Fusion
• HtSeq
• RSeQC
• GATK
• UCSC Genome Browser

Full Length cDNA Sequencing (mRNA- Seq) results delivery format (Appendix)

Advancing scientific research is a primary motivation of all bioinformaticians and acknowledgment of contribution through authorship on manuscripts arising from this analysis is one way our work is assessed and attributed. We request to be considered for authorship on any manuscripts using the analysis results provided if you believe we have made substantive intellectual contributions to the study.