Next Generation Sequencing (NGS)/Expression quantification

RNA-seq expression quantification

Overview

 * Objective: to estimate the relative abundance of transcripts from an rna-seq sample.
 * Biological questions?


 * Inputs:
 * Reference:
 * Annotated Genome (fasta/gff)
 * Transcriptome (fasta)
 * RNA-seq raw reads (fastq, Ilumina).


 * Outputs:
 * Relative abundance per transcript.

Experimental Design
See (link) RNA sequencing

Typical Steps in the Method: (table or workflow or paragraph?) - Quantification | Get transcript abundance |
 * Data generation | Get some data. | Seq (link) Sequencing RNA technologies,
 * Get a reference | You need a reference with all the involved transcripts for which you want relative abundance |
 * RNA-seq read QC | Check the quality of the sequencing itself. |
 * Adaptor trimming / quality filtering / contamination filtering | Reads will contain sequence that is not from the transcript but from the technology. |
 * Mapping | Align your reads to the reference |

Real life workflows
- - - - -

Command line scripts
Example workflow:

Real life workflows: - - - - -

Some tools and resources

 * Get a reference: MetaDB (link)genome assembly and annotation (link) transcriptome denovo assembly ....
 * QC: SeqWiki: FastQC, KAT
 * Trimming: SeqWiki; Trimmomatic, Sickle, Cutadapt....
 * Mapping: Seqwiki: tools...... ... ... ... ... ... ... .. ....
 * Quant: Seqwiki: tools