Application of next-generation sequencing for comparative transcriptome analysis

I have used novel whole transcriptome sequence data generated from massively parallel high-throughput next generation sequencing technologies, namely 454 pyrosequencing and Illumina sequencing, to perform comparative transcriptome analyses of C. elegans populations in specific biological conditions and developmental stages. Firstly, I have conducted transcriptome profiling of C. elegans in its first larval (L1) stage using data generated from the Roche 454 sequencing platform. I have used this data to refine gene structures, identify putative novel transcripts, and characterize the L1 specific transcriptome. Secondly, I have taken Illumina paired-end sequencing transcriptome data generated from starved and fed populations of C. elegans in the L1 stage and investigated metabolic and developmental gene expression changes that are specific to this developmental stage. Additionally I have taken advantage of this paired-end data to perform de novo assembly of the short tag sequences to identify novel transcript structures with confirmed splice sites, providing greater confidence in novel gene structure discovery. Lastly, I have investigated global transcriptional responses to oxidative stress in aging defective aak-2 mutants. This analysis has led to the identification of potential downstream targets of the AMP-activated protein kinase, AAK-2, which is known to be involved in stress resistance and lifespan control in C. elegans. This study demonstrates the impact of new high-throughput sequencing technologies on genomic analysis, providing valuable insight into transcriptome complexity as a result of developmental, metabolic, and mutational effects in C. elegans. The approaches I have described here can be applied to transcriptome analyses of more complex organisms ultimately leading to investigation of transcriptional changes in human diseases.