The Genome Sequencer 20™ System, developed by 454 Life Sciences, is an ultra-high-throughput automated DNA sequencing system capable of resolving hundreds of thousands of DNA sequences in one run. The basic chemistry utilizes the release of pyrophosphate (PPi) that occurs with each nucleotide addition during DNA-directed DNA synthesis to generate an amount of light commensurate with the amount of PPi released; this light is captured by a CCD camera and converted into a digital signal. The combination of signal intensity and positional information over the PicoTiterPlate™ device (see below) allows the Sequencer's Linux-based computer, equipped with an onboard Field Programmable Gate Array processor, to determine the sequence of hundreds of thousands of individual reactions simultaneously, producing millions of nucleotides of sequence per hour.
The Genome Sequencer 20 System provides a complete solution for ultra-high-throughput DNA sequencing. The system includes the Genome Sequencer 20 Instrument and accessories; software to generate basecalls and Phred-base quality scores for raw reads ; software to assemble the overlapping reads de novo into a consensus genome; software to map the reads to a known genome, and consumable kits required for library construction, clonal amplification, and sequencing.
Software
Run Training: the individual read output is filtered for quality, and basecalls and Phred-equivalent quality scores are generated for each base within each filtered read. Sequence is output as a FASTA file with an associated file of Phred-equivalent quality scores.
Mapping: The reads are mapped to a FASTA reference genome provided by the user. Signals from overlapping reads are combined and processed into contigs, generating a consensus for each basecall that is typically more accurate than basecalls from the individual reads. High confidence differences from the FASTA reference genome are identified. Contigs are output as a FASTA file with an associated file of Phred-equivalent quality scores.
Assembly: Reads are filtered for quality, and basecalls are generated for each read. Reads are processed to find overlapping regions, and assembled into contigs. Basecalls are generated from the consensus of the overlapping reads. Contigs are output as a FASTA file with an associated file of Phred-equivalent quality scores.
