Next Generation Sequencing and Bioinformatics
The GenePool operates two Illumina HiSeq 2000 sequencing systems, and two MiSeq systems.
Each HiSeq is capable of generating tens of gigabases per day. These data can be...
The instrument are run using a 'flowcell' that has 8 lanes. We can run a different sample in each lane or multiplex several samples per lane. So whether you want a "small" number of short reads or hundreds of gigabases, we can process your samples...
The Illumina sequencing platforms use a small flowcell to immobilise, amplify and sequence up to 1.5 billion molecules at once. HiSeq flowcells are split into eight lanes, allowing 8 independent experiments (reading from 180 million clonally amplified fragments each) in a single run. MiSeq flowcells currently use a single lane.
Read length on the MiSeq is up to 150 bases.
On the HiSeq 2000 platform we support up to 100 base reads.
Paired end and mate pair reads
The technology also allows paired-end sequencing (e.g. the sequencing of 100 bases from each end of a DNA fragment). Thus a single paired-end run can yield over 300 gigabases of DNA sequence. These paired reads can be used to generate better genome assemblies, map alternative transcripts/splicing, and also to index samples mixed into one flow cell lane. We can also construct mate pair libraries for sequencing, where the two reads that are generated derive from positions several kilobases apart in the original genome.
Please see our Delivery Goals for more details on guaranteed throughput for each platform.
The Illumina platform is ideal for resequencing of genomes to detect changes from a reference sequence. For bacterial genomes, Illumina resequencing improves on genome chip hybridisation in that it can detect and assemble segments of DNA not present in the reference. For larger genomes Illumina is ideal for detection of mutations (SNPs and indels) in individual organisms or inbred lines.
One Illumina HiSeq 200 lane can yield 8,000-fold coverage of an E. coli, 3,000-fold coverage of a S. cerevisiae, 360-fold coverage of a C. elegans, 200-fold coverage of a D. melanogaster and 12-fold coverage of a human genome. The platform is also suited to directed or targeted resequencing (where a smaller portion of a genome is PCR amplified and sequenced). To make best use of this capacity, individual samples can be indexed and mixed on a single lane.
De novo genome sequencing
While 50 base reads do not at first glance appear to be a very useful for genome sequencing, 50 bases is long enough to be unique in over 90% of the human genome (3 Gb) and 96% of the C. elegans genome (100 Mb).
Assemblers are already available for the de novo generation of small genome sequences, and with the availability of paired-end and mate-pair reads, even larger genomes are accessible to direct sequencing with the Illumina HiSeq platform. We can generate data for a first draft de novo sequencing of bacterial and other genomes up to ~ 1 Gb at present (we recommend at least 60 fold over sequencing with paired-end and mate-pair reads), and can assemble these to yield a draft assembly. Depending on the genome sequenced, this assembly may be in as few as tens of contigs, but can be up to tens of thousands. The draft is good for gene prediction, gene finding, and as a platform for directed finishing. Paired-end reads improve these assembly statistics significantly. For genome sequencing de novo, the recommendation is to use a mix of paired-end and mate-pair libraries to generate longer contigs.
Digital transcriptomics and ChIP-sequencing
Using an approach akin to serial analysis of gene expression (SAGE), the Illumina platforms can specifically capture and sequence 3' tags from mRNA populations. These tags can easily be sorted, and for a sequenced transcriptome assigned to genes, to provide a digital transcriptome for a cell line, tissue or organism. These digital transcriptomic datasets have a dynamic range of ~4 orders of magnitude, with essentially no need for 'background correction' and thus are likely to be superior than conventional microarrays for many applications. A similar application of cDNA sequencing (called RNA-Seq in the literature) can be used to map and count transcripts and alternative transcripts.
Similarly, the Illumina HiSeq platform can be used to assess the sequence diversity in, and thus relative occupancy of, DNA or RNA sites bound by specific regulators or modified in different ways, using a ChIP-sequencing approach.
For more information, including pricing, contact us.
|The GenePool has core support from:||GenePool admin site|
|The University of Edinburgh
School of Biological Sciences
The Darwin Trust