Whole Transcriptome Shotgun Sequencing (WTSS) Data Analysis

Whereas microarrays are important in the identification and quantification of the mRNA transcripts, they are encumbered by the inability to identify novel transcripts, limited dynamic range for detection, and difficulty in replicability and inter-experimental comparison. RNA Seq, also known as whole transcriptome shotgun sequencing (WTSS) overcomes many of these problems. Making use of high-throughput next-generation sequencing methods, sequencing the entire transcriptome permits both transcript discovery and robust digital quantitative analysis of gene expression levels.

To achieve its two major functions (transcript discovery and gene expression quantification), WTSS relies on the generation of short reads of transcript sequence information. Its advantages include high-throughput sequencing, lack of bacterial cloning constraints, high coverage, ability to discover new exons and handle RNA splicing, and low cost. This course equips students with the following skills:

• Poly (A)-selection of RNA
• Fragmentation of RNA to an average length
• Conversion into cDNA
• Sequencing.
• Mapping of the reads onto the genome
• Calculation of the transcript prevalence

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