Here is my second summary from 4th NGS Congress at London Heathrow end of 2012. It will bring to you some (hopefully) interesting new facts about sequencing with PacBio RS – the second long read technology present in the actual markets and also the only system delivering reads even longer than 10,000 bp…
Kevin Corcoran, Senior Vice President at Pacific Biosciences held an interesting and very nice talk about the most recent developments for the PacBio RS system. He also showed some nice detailed road maps about future aims and plans. One important thing actual to be mentioned is the launch of the new “XL Chemistry” – while still “C2 Chemistry” may be used as well. The other very interesting story is about “Stage Start” a new feature enabling a parallel start of all sequencing detection similar to the well-known “hot start” technology for PCR. Such detection of sequences better will start from a defined position for most of the libraries than starting from somewhere in the middle. Last but not least, I’m very keen to learn how the future “Photo Protected DNA Polymerases” may further develop – an idea being really very, very next-next-generation…
First of all I can summarize that applying “XL Chemistry” looks really interesting and this being true also in terms of Eurofins MWG Operon de novo sequencing and assembly focus. This new feature of the PacBio RS machine may also open some new doors to other types of applications, while in general the need for extrem high data coverages may be reduced in parallel.
Currently “C2 Chemistry” is on the machine and running a 90 min video may deliver you about 20-50,000 reads and data outputs of 30-50 Mb – of course higher yields may be possible for “ideal” DNA samples. The average read length is about 3,000 bp (!), while the 95% percentile is about 8,000bp. With the new “XL Chemistry” we got an average yield of about 40,000 reads per SMRT cell with an average read length of about 4,000bp (+30%). Overall, we are very pleased with these first results, especially since we see some good potential to further increase data yields using the new software pipeline started in parallel (Hierarchical Genome Assembly Process and Quiver).
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It is also important to mention two different ways of “How-to-deal” with the XL Chemistry. 1) “XL chemistry for Polymerase binding”, but “C2 chemistry for sequencing”. This allows for longer reads at the same quality (currently we still do have a single error rate of 10% to 20 %, average maybe 15%). 2) “XL chemistry for Polymerase binding” AND “XL chemistry for sequencing”. Such one can yield even longer reads, but unfortunately the error rate will also increase by a few %. Therefore this method is being recommended especially for de novo assembly or finishing genomes.
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Finally one real “next-next-gen” highlight was the presentation of a development at Pacific Biosciences scoping with the idea to protect the polymerase enzyme from being killed by the energy of the laser. A picture shows how this should work in principle – by setting in place a laser-light protecting sun-blocker – this story was really fascinating for me and I hope to see in future more than the very promising first data results …
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So over all Pacific BioSciences keeps also moving very fast in year 2013 and it will be very nice to see and learn how all these additional improvements and new features may improve the overall data results of this fascinating very long read technology offering today real single reads longer than 10,000 bp.
Cheers now and see you on our next BLOG,