Editing DNA sequencer (section)

===Roche===

The 454 DNA sequencer was the first next-generation sequencer to become commercially successful.<ref name="Comparison Next-Gen Seq">{{cite journal |author1=Lin Liu |author2=Yinhu Li |author3=Siliang Li |author4=Ni Hu |author5=Yimin He |author6=Ray Pong |author7=Danni Lin |author8=Lihua Lu |author9=Maggie Law |title = Comparison of Next-Generation Sequencing Systems|journal = Journal of Biomedicine and Biotechnology|year=2012 |volume = 2012|pages = 251364|pmid = 22829749|doi = 10.1155/2012/251364|pmc=3398667 |doi-access=free }}</ref> It was developed by [[454 Life Sciences]] and purchased by Roche in 2007. 454 utilizes the detection of pyrophosphate released by the DNA polymerase reaction when adding a nucleotide to the template strain.

Roche currently manufactures two systems based on their pyrosequencing technology: the GS FLX+ and the GS Junior System.<ref>{{Cite web |url=http://454.com/products/index.asp |title=Products : 454 Life Sciences, a Roche Company<!-- Bot generated title --> |access-date=2012-09-05 |archive-url=https://web.archive.org/web/20120913054317/http://454.com/products/index.asp |archive-date=2012-09-13 |url-status=dead }}</ref> The GS FLX+ System promises read lengths of approximately 1000 base pairs while the GS Junior System promises 400 base pair reads.<ref>{{Cite web |url=http://454.com/products/gs-flx-system/index.asp |title=Products - GS FLX+ System : 454 Life Sciences, a Roche Company<!-- Bot generated title --> |access-date=2012-09-05 |archive-url=https://web.archive.org/web/20120905145227/http://454.com/products/gs-flx-system/index.asp |archive-date=2012-09-05 |url-status=dead }}</ref><ref>{{Cite web |url=http://454.com/products/gs-junior-system/index.asp |title=Products - GS Junior System : 454 Life Sciences, a Roche Company<!-- Bot generated title --> |access-date=2012-09-05 |archive-url=https://web.archive.org/web/20120913065444/http://454.com/products/gs-junior-system/index.asp |archive-date=2012-09-13 |url-status=dead }}</ref> A predecessor to GS FLX+, the 454 GS FLX Titanium system was released in 2008, achieving an output of 0.7G of data per run, with 99.9% accuracy after quality filter, and a read length of up to 700bp. In 2009, Roche launched the GS Junior, a bench top version of the 454 sequencer with read length up to 400bp, and simplified library preparation and data processing.

One of the advantages of 454 systems is their running speed. Manpower can be reduced with automation of library preparation and semi-automation of emulsion PCR. A disadvantage of the 454 system is that it is prone to errors when estimating the number of bases in a long string of identical nucleotides. This is referred to as a homopolymer error and occurs when there are 6 or more identical bases in row.<ref>{{cite journal|last=Mardis|first=Elaine R.|s2cid=2484571|title=Next-Generation DNA Sequencing Methods|journal=Annual Review of Genomics and Human Genetics|date=1 September 2008|volume=9|issue=1|pages=387–402|doi=10.1146/annurev.genom.9.081307.164359|pmid=18576944}}</ref> Another disadvantage is that the price of reagents is relatively more expensive compared with other next-generation sequencers.

In 2013 Roche announced that they would be shutting down development of 454 technology and phasing out 454 machines completely in 2016 when its technology became noncompetitive.<ref>{{Cite web |date=2013-10-15 |title=Roche Shutting Down 454 Sequencing Business |url=https://www.genomeweb.com/sequencing/roche-shutting-down-454-sequencing-business |website=GenomeWeb |language=en}}</ref><ref>{{Cite web |last=Davies |first=Kevin |date=April 23, 2013 |title=Roche Shuts Down Third-Generation NGS Research Programs |url=https://www.bio-itworld.com/news/2013/04/23/roche-shuts-down-third-generation-ngs-research-programs |website=Pubs - Bio-IT World |language=en}}</ref>

Roche produces a number of software tools which are optimised for the analysis of 454 sequencing data.<ref name=454analysis>{{Cite web |url=http://454.com/products-solutions/analysis-tools/gs-de-novo-assembler.asp |title=Products - Analysis Software : 454 Life Science, a Roche Company |access-date=2013-10-23 |archive-url=https://web.archive.org/web/20090219223046/http://454.com/products-solutions/analysis-tools/gs-de-novo-assembler.asp |archive-date=2009-02-19 |url-status=dead }}</ref> Such as,

* ''GS Run Processor'' converts raw images generated by a sequencing run into intensity values.<ref>[http://sequence.otago.ac.nz/download/ManualPartB.pdf Genome Sequencer FLX System Software Manual, version 2.3]</ref> The process consists of two main steps: image processing and signal processing. The software also applies normalization, signal correction, base-calling and quality scores for individual reads. The software outputs data in Standard Flowgram Format (or SFF) files to be used in data analysis applications (GS De Novo Assembler, GS Reference Mapper or GS Amplicon Variant Analyzer). 
* ''GS De Novo Assembler'' is a tool for ''de novo'' assembly of whole-genomes up to 3GB in size from shotgun reads alone or combined with paired end data generated by 454 sequencers. It also supports de novo assembly of transcripts (including analysis), and also isoform variant detection.<ref name="454analysis" /> 
* ''GS Reference Mapper'' maps short reads to a reference genome, generating a consensus sequence. The software is able to generate output files for assessment, indicating insertions, deletions and SNPs. Can handle large and complex genomes of any size.<ref name="454analysis" /> 
* Finally, the ''GS Amplicon Variant Analyzer'' aligns reads from amplicon samples against a reference, identifying variants (linked or not) and their frequencies. It can also be used to detect unknown and low-frequency variants. It includes graphical tools for analysis of alignments.<ref name="454analysis" />