[mira_talk] Re: Problem with Paired 454, MP Illumina, MIRA and Bambus

  • From: Nestor Zaburannyi <nestor@xxxxxxxxxxxxx>
  • To: Surya Saha <mira_talk@xxxxxxxxxxxxx>
  • Date: Fri, 15 Jul 2011 18:46:07 +0200

Dear Surya,

> Dear Nestor,

> I have used them only with Illumina data but I don't recall that they cannot
> be used with 454 paired reads. Is it explicitly mentioned anywhere?

In this topic there is mentioning of 454:
http://seqanswers.com/forums/showthread.php?t=8350

Apart from that, SSPACE is using bowtie with options that exclude any reads 
even with one mismatch. And this is hardly the truth for 454. Especially 
unclipped, raw data.


> In case you are using Velvet, SOPRA can be run in integration with Velvet
> and the authors suggest that is better than running SOPRA separately
> post-assembly although I have not tried using it this way (yet). Good luck!

> -Surya

> On Fri, Jul 15, 2011 at 12:21 PM, Nestor Zaburannyi 
> <nestor@xxxxxxxxxxxxx>wrote:

>> Dear Surya,

>> > Hi Nestor,

>> > I have had moderate luck with two scaffolders besides the ones already
>> > mentioned. You might want to try them out.

>> > SSPACE (
>> >
>> http://www.baseclear.com/sequencing/data-analysis/bioinformatics-tools/sspace/
>> > )
>> > SOPRA (http://www.physics.rutgers.edu/~anirvans/SOPRA/)

>> > -Surya

>> Thanks, already gave SSPACE a try. It is good, pity only for short reads as
>> i understand. Will also try also SOPRA.


>> > 2011/7/12 Nestor Zaburannyi <nestor@xxxxxxxxxxxxx>

>> >> Dear Bastien,

>> >> Thanks for your hints. Indeed, dirty hack involving the reversal of both
>> MP
>> >> read sets helped.

>> >> Regards
>> >> Nestor

>> >> > On Jul 6, 2011, at 23:19 , Nestor Zaburannyi wrote:
>> >> >> I have 454 paired data and Illumina Mate-Pair data. It took me some
>> time
>> >> to figure out that not the Bambus is wrong... Long story short:

>> >> > Actually I am not so sure that Bambus is correct :-)

>> >> > One thing to keep in mind are the orientation of the pairs and what
>> the
>> >> scaffolder expects:
>> >> > - Sanger pairs are oriented like this:   ------>    <--------  and
>> that
>> >> is what Bambus wants per default
>> >> > - 454 pairs are originally oriented like this:  -------->
>>  --------->
>> >> but as scaffolders (and MIRA) originally did not expect that, I use a
>> trick
>> >> by letting sff_extract reverse one sequence so that everything as back
>> to
>> >> "normal". Nowadays MIRA could also use the forward / forward
>> orientation,
>> >> but I had not time to change sff_extract.
>> >> > - Illumina paired-end reads look like this:    -------->   <---------
>> >>  ... which makes it easy.
>> >> > - Illumina mate-pairs look like this:    <---------    ---------->
>>  ...
>> >> which again lets scaffolders like Bambus despair as the expect something
>> >> different. MIRA does not care there, it can work with that, too.

>> >> >> Most of the contigs are not joined properly and we get hundreds of
>> >> thousands "Invalid" links with consistent distance between them.
>> Something
>> >> like:

>> >> > Which is a symptom of what I described: mate-pairs are facing
>> outwards,
>> >> Bambus expects them to face inwards.

>> >> > There is no easy solution to this, the simplest one I can think of is
>> to
>> >> reverse all reads of the Illumina mate-pair set (both sets!) and then
>> >> assemble. Should keep Bambus happy.

>> >> > Hope that helps,
>> >> >   Bastien






>> >> --
>> >> З повагою,
>> >>  Nestor                            mailto:nestor@xxxxxxxxxxxxx


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>>  Nestor                            mailto:nestor@xxxxxxxxxxxxx


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З повагою,
 Nestor                            mailto:nestor@xxxxxxxxxxxxx


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