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Roche 454 Sequencing Background

Roche 454 Sequencing Background

454 Sequencing is a Next-Generation Sequencing, or NGS, technology.  Data is generated using pyrosequencing chemistry where clonally amplified libraries are sequenced by synthesis.  Libraries are prepared using emulsion-based PCR (emPCR) amplification on beads.  Each bead occupies a well on a Pico-Titer-Plate or PTP plate and the nucleotides in the sequence are determined by a chemiluminescent signal occurring during nucleotide incorporation.  This signal is captured by a high resolution camera and is proportional to the number of nucleotides incorporated.  The signals are processed and sequence is determined.

A single instrument run produces up to 1 million sequence reads that are on average 400bp, and will soon approach 800bp.  For additional flexibility, runs can be multiplexed bioinformatically through the use of barcoding or MID (Multiplex Identifiers) adapters, or physically through the use of 2, 4, 8 or 16-region gaskets.

The throughput and read lengths offered by this system make it an ideal platform for:

  • Whole-genome sequencing of small or medium genomes
  • Amplicon Sequencing
  • Transcriptome sequencing
  • Targeted Resequencing
  • Metagenomics

454 sequencing is an established NGS technology that has been cited in over 1,500 papers since 2005.  It is an ideal platform for many projects where read length is important.   Additional background information about the technology behind 454 sequencing can be found here.

For more information on this service and its applications to your research, visit our Roche 454 Sequencing Protocols and Resources page.

 

Page last updated March 18, 2012 @ 5:25 pm