The Roche/454 Life Sciences GS FLX and GS Junior: an obituary

You were a pioneer, the first successful ‘next generation’ (if you’ll pardon the term) commercially available sequencing platform in 2005. You just beat Solexa, but it was a fairly close call.

gsflx-uio1

The author (left) with colleagues showing off their 454 GS FLX

Your greatest accomplishment was to show that pyrosequencing, which was around for a while already, could be scaled up, both in terms of read length and parallelisation. You started the revolution in DNA sequencing, suddenly making large scale genomic projects available to labs that traditionally only could dream of been doing such projects at this scale.

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Make Newbler open source: the Roche response and the future of Newbler

Earlier this year, I started a petition to ask Roche/454 Life Sciences to make the Newbler software (gsAssembly, gsMapper and Amplicon Variant Analyzer) open source. See this post for the background to the petition.

Source: Wikimedia Commons, by Marcus Quigmire

Source: Wikimedia Commons, by Marcus Quigmire

When I closed the petition, 162 people had signed it, see the PDF on figshare. During the Advances in Genome Biology and Technology (AGBT) meeting in Florida, I handed over the results of the petition to two Roche representatives, Dan Zabrowski, Head of Roche Sequencing Unit and Paul Schaffer, Vice President of Roche 454 Sequencing Business, see my blog post on the conversation I had with them.

Dan Zabrowski and Paul Schaffer promised me an official Roche response, and here it is (exclusively released through this blog): Continue reading

Make Newbler open source: petition results and the meeting with Roche

A couple of weeks ago, I started a petition to ask Roche/454 Life Sciences to make the Newbler software (gsAssembly, gsMapper and Amplicon Variant Analyzer) open source. See this post for the background to the petition.

The results are in, see the PDF on figshare. 162 people have signed the petition. Many thanks to all of you!

This week, I attended the Advances in Genome Biology and Technology (AGBT) meeting in Florida, and on Thursday I handed over the results of the petition to two Roche representatives, Dan Zabrowski, Head of Roche Sequencing Unit and Paul Schaffer, Vice President of Roche 454 Sequencing Business.

I had a half hour, very open and interesting discussion with Roche. Roche expressed their appreciation of the fact that we as a community voiced our concerns and wishes around Newbler. Roche occasionally picks up signals from researchers, but a petition like this was very useful for them as a much stronger signal of what we think about one of their products.

Dan Zabrowski told me Roche is committed to fully support access to the Newbler software even after the 454 Life Sciences shutdown. They will take the request for open source access to the code seriously, and promised to come with an official response somewhere in the coming weeks. They did not hint at what that response would be, which is understandable.

I want to thank Dan Zabrowski and Paul Schaffer for giving me time to explain the background and hand them the results. I also again want to thank all of you who signed the petition. We may collectively have made a difference. Keep an eye out on my twitter feed and this blog for the official Roche response!

Make Newbler Open Source: an update

My petition to ask Roche/454 Life Science to make the Newbler software open source, as announced on this blog, has so far yielded 124 signatures. Thanks to all who signed!

It is not to late to add your signature!

Please do this before Thursday February 13th. On that day, at the Advances in Genome Biology and Technology (AGBT) meeting, I have been given the opportunity to hand over the petition results to two Roche representatives: Dan Zabrowski, Head of Roche Sequencing Unit and Paul Schaffer, Vice President of Roche 454 Sequencing Business. The fact that Roche opens up for me meeting these people makes me very happy!

Finally, here are some of the comments added by people who signed the petition. I anonymised them, but if you see your comment and would rather not I repeat it here, let me know (lex dot nederbragt at ibv dot uio dot no). Thanks for these wonderful comments on the Newbler assembly:

Roche has an excellent opportunity of publicit their commitment with science making their fantastic product Newbler open-source

I would like to runProject forever!

Newbler source code is very valuable to the sequencing community. Roche will be doing a service to the scientific community by making the Newbler source code available after the 454 shutdown.
Goodwill from making this OS is best value Roche can get for this code
This is way overdue. James Knight has done wonderful work but Newbler may fall through the cracks if it is not opened up to the community.

The newbler is great assembler for paired end data.
Please make it opensource, and I would be happy to contribute to it’s development.

Open source is a friend to science aiding reproducibility and allowing future developers to learn from existing work. Please do not limit science by preventing our access.

The Newbler suite has been remarkably useful and should continue to be so. I hope Jim Knight sees this and pushes for his creation to be made open source

I strongly support this petition and note there is a precedent for a commercially-developed assembler to be made publicly available under an open source license – the Celera assembler – much to the benefit of the scientific community.

Newbler was the basis for contig assembly of MiSeq data for me. An excellent piece of software.

Newbler was always my favorite thing about 454. It deserves to live on.

Make Newbler open source

(Cross-posted at contig.wordpress.com)

Typical newbler assembly output

Typical Newbler assembly output

The Newbler assembler and mapper (gsAssembler, gsMapper) was developed especially for working with the reads from the Roche/454 Life Science sequencing technology. It is one of the best programs to deal with this type of data, scoring well in the assemblathon 2 competition. Newbler has been used for many large and small genome assemblies (numerous bacteria, Atlantic cod, bonobo, tomato, to name a few). Recently, Newbler has added support for using multiple sequencing technologies, making it one of the few hybrid assembly programs available. At the Advances in Genome Biology and Technology (AGBT) in 2013, Roche announced having used the Newbler program with a hybrid 454 and Illumina dataset to improve upon the human genome.

However, the Newbler program is not open source. Luckily, researchers only need to fill out an online form to get a free copy of the software. Still, this has hampered the wide-spread adoption of this program. Newbler, for example, was not included in assembly evaluations like GAGE and GAGE-B. That Roche/454 does not want to make the source code for Newbler available is partly understandable from a commercial standpoint: at least one competitor technology (Life Tech/Ion Torrent) with a similar sequencing error-model could benefit from access to the code. In fact, in a blog post, I showed Newbler to be superior to an open-source program when assembling Ion Torrent mate-pair data.

More worringly is that the hundreds of projects that used Newbler as part of the analysis are fundamentally irreproducible without the source code for each of the different versions. This is especially the case for projects, such as the Atlantic cod genome project, that have been given access to development versions of the code, incorporating elements not available to the general community.

Last October, Roche announced it will shutdown its 454 sequencing business in mid-2016. Whatever one may feel about this decision, this further strengthens the argument for Roche/454 to make the Newbler source code open source. After the 454 shutdown, Newbler is otherwise likely to disappear too, meaning that large swathes of the literature cannot be recapitulated from the raw data. Also, long after the 454 shutdown, many researchers will have to process their 454 sequencing data, and many may still want to rely on Newbler for that purpose.

There are several other reasons why I feel the research community should be given access to the source code of Newbler. Newbler represents a very valuable contribution to the field of genome assembly and mapping. Software developers can learn from the algorithms and implementations of the Newbler code, opening up for reusing these in other programs. Also, there is the hope that developers will improve upon the program, for example by adding support for other sequencing technologies, or assembling with reads longer than the current maximum of 2 kbp.

So I hereby ask the readers of this blog for help: I have set up an online petition asking for Roche/454 to make the Newbler source code available at the latest at the time of the 454 shutdown. Please sign the petition here. Additionally, spread the word (e.g., on twitter or your own blog). Thanks in advance!

I intend to hand over the results of the petition to a Roche representative at the Advances in Genome Biology and Technology (AGBT) meeting (February 12-15, 2014).

Finally, feel free to use the comments to tell me about your Newbler experiences!

(Thanks to Nick Loman for his constructive comments on an earlier version of this post)

Fast genome sequencing of pathogenic bacteria – which benchtop instrument to choose?

Nick Loman was kind enough to give me an advanced copy of his paper in Nature Biotechnology entitled “Performance comparison of benchtop high-throughput sequencing platforms” (Loman et al, 2012). I thought to present a quick summary of the paper here and add some comments of my own.

The paper sets out to “compare the performance of three sequencing platforms [Roche GS Junior, Ion Torrent PGM and Illumina MiSeq] by analysing data with commonly used assembly and analysis pipelines.” To do this, they chose a strain from the outbreak of food-borne illness caused by Shiga-toxin-producing E. coli O104:H4, which caused a lot of trouble in Germany about a year ago. The study is unique in that it is focuses on the use of these instruments for de novo sequencing, not resequencing.

First, they used the ‘big brother’ of the GS Junior, the GS FLX, to generate a reference genome (combining long reads obtained using the GS FLX+, and mate pairs using Titanium chemistry). Then, the same strains were sequenced on the benchtop instruments, and these reads were compared to the reference assembly. The reads were both compared directly, and after assembly with a few commonly used programs.

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Ion Torrent Mate Pairs and a single scaffold for E coli K12 substr. MG1655

(The impatient reader might want to skip to the conclusion at the end of this post…)

Last wednesday, Ion Torrent released a tech note and associated run data with shotgun (single-end) and Mate Pair runs for Escherichia coli K12, substrain MG1655. Both a 3.5 kb and 8.9 kbp insert size, as well as a shotgun library, were sequenced on a 316 chip each. In the tech note, they describe assemblies using different combinations of the data, and show how adding the mate pairs yields assemblies with fewer scaffolds and gaps. The Ion mate-pair protocol is very similar to the one used by 454 Life Sciences for their (unfortunately called) Paired-end libraries: long fragments are circularized using a linker sequence, and sequencing is peformed across this linker, allowing for easy identification of the pair halves.

This is the first real ‘long-distance’ Mate Pair data from Ion Torrent, which is exciting and made me have a close look at it. I was especially interested in how the newbler program, developed by 454 Life Science for their 454 reads, would perform on these data.

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