My talk will be a summary of my research so far in investigating the effects of artefacts which occur during the library preparation stage of Next generation sequencing.
Next generation sequencing (NGS) of DNA has dramatically transformed approaches to genomic and genetic research. DNA sequencing refers to a laboratory method used to determine the sequence of a DNA molecule. Some of the well-known technologies that are applied in this process include the Roche GS-FLX 454 Genome Sequencer (originally 454 sequencing), the Illumina Genome Analyser (originally Solexa technology), the ABI SOLiD analyser, Polonator G.007 and the Helicos HeliScope platforms. These technologies (also referred to as massively parallel sequencing technologies) have enabled the sequencing of DNA at unprecedented speeds compared to the "original" sequencing methodology known as the Sanger method.
Although NGS has revolutionised biology by increasing current understanding of many genes and genomic regions involved in the pathogenesis of human diseases, there are challenges associated with the use of these new technologies. For example, the sequencing of parts of a genome characterized by extremely biased base composition is still a great challenge to the currently available NGS platforms. The genomes of certain important pathogenic organisms like Plasmodium falciparum and Escherichia coli display extremes of base composition, high AT content and high GC content respectively. The sequencing "coverage" of these genomes can be affected by artefacts that may be introduced at various stages of the sequencing process, thus affecting final sequencing output.
My project studies the effects of artefacts that occur during the initial stage of the sequencing work flow referred to as library preparation. The purpose of this research is to identify known artefacts that can occur during the library preparation stage from existing literature, and to analyse the extent to which sequencing coverage may be affected by these artefacts. In other words, a study of problems that can arise from the library preparation stage of the sequencing work flow and how they affect final sequencing output forms the basis of my work.
- Oyola, S.O., Otto, T.D., Gu, Y., Maslen, G., et al. (2012) Optimizing Illumina next-generation sequencing library preparation for extremely AT-biased genomes. BMC genomics.
- Sanger, F., Nicklen, S. & Coulson, A.R. (1977) DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences of the United States of America.
- Zhang, J., Chiodini, R., Badr, A. & Zhang, G. (2011) The impact of next-generation sequencing on genomics. Journal of genetics and genomics = Yi chuan xue bao.