[mira_talk] Re: PacBio for scaffolding?

  • From: 000.calabi.yau.000@xxxxxxxxxxxxxx
  • To: mira_talk@xxxxxxxxxxxxx
  • Date: Tue, 19 Jul 2011 20:29:05 +0200

I wonder if they actually CAN improve the single pass accuracy much more. I have the feeling that with single molecule sequencing you are close to a physical barrier of what is possible as compared to second gen sequencers where you average your signal over a clonal population. I think Helicos data also has problems with dark bases. The closer you look the noisier the data i guess... And as I understood it, every time you read sequence you zap some nucleotides with the laser in the soup and bleach them. If my understanding is right... every time you look at a base (i.e. sequencing) you increase the chance that the following sequence will have an error. Is that actually true? Plus if the polymerase incorporates the wrong base it also pops up.


Of course with consensus sequencing you get a better picture. But then I wonder if the throughput is enough...

But I find long reads and/or strobed reads much more interesting because they open some really cool possibilities that other chemistries can't really offer (yet)... one has to see what will come out of this..

Thanks for all the input. I don't really have many people to talk to around here. :-)


Cheers,

Markus




On Tue, 19 Jul 2011 10:15:14 +0200, Brian Forde <bforde@xxxxxxxxx> wrote:

Did PacBio not recently announce that they had increased read lengths to an average of 2.9kb and with consensus circular sequencing increased accuracy
to 99.998%. I suppose the question then is whether consensus circular
sequencing significantly increases the cost of a project or not.

Brian

On Mon, Jul 18, 2011 at 11:54 PM, Bastien Chevreux <bach@xxxxxxxxxxxx>wrote:

On Jul 18, 2011, at 23:25 , Robin Kramer wrote:
> I doubt that pacbio is going to bring much to the assembly world until
that accuracy improves.  The problem is the reads will still need to be
"error corrected" by competing sequencing technologies(similar to the
problem that 454 reads have),

Math is simple: if PB gets reads >= 1500 at >=95 %, 454 unpaired reads are
in danger of extinction.

I don't think that this is out of reach.

Bacteria and lower eukaryotes would then have a mix of PB CCS, 454 PE and Solexa. Or perhaps even PB CCS, PB strobed abd Solexa ... but for that the
single read accuracy of Solexa should go >= 90%.

B.


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