Wayne:High Throughput Sequencing Resources

High throughput (HT) platform and read types

• Illumina single-end vs. paired-end
• 454 Roche
• SOLiD
• MiSeq
• Ion Torrent

File formats and conversions

• bcl
• qseq
• fastq

Deplexing using barcoded sequence tags

• Editing (or hamming) distance

Quality control

• Fastx tools
• Using mapping as the quality control for reads

Trimming and clipping

• Trim based on low quality scored per nucleotide position within a read
• Clip sequence artefacts (e.g. adapters, primers)

RNA-seq analysis

• Quantifying and annotating aligned reads
• DESeq
• edgeR

A variety of additional R packages are available for normalizing RNA-Seq read count data and identifying differentially expressed genes (DEG):

• easyRNASeq (simplifies read counting per genome feature)
• DEXSeq (Inference of differential exon usage)
• DEGseq
• baySeq (also see: segmentSeq)
• Genominator (Bullard et al. 2010)

Basic server commands (for Sirius)

Command	Usage
pwd	Print working directory (your current location
ls	List (all contents of current location)
ls options	ls -a (hidden files), ls -l (long/detailed list), ls -t (sorted by time modified instead of name)
cd	Change directories
mkdir	Make directories
head filename	Print to screen the top 10 lines or so of the specified file
tail filename	Print to screen the last 10 lines or so of the specified file
more filename	Allows file contents or piped output to be sent to the screen one page at a time
less filename	Opposite of more command
whereis [filename, command, etc]	Lists all occurances of filename or command
mv	Move (akin to cut/paste), to remove the file in the current location; Usage: mv current/path/filename destination/path/filename
cp	Copy (also used to rename files if you keep them in their current path), keeps a copy in the current path; Usage: cp current/path/filename destination/path/filename
1.5-2μL	0.8μL
1μL	0.4μL
1μL	0.4μL
0.1μL	0.08μL
1μL	1μL

HT sequence analysis using R (and Bioconductor)

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