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Forum: Bioinformatics 08-20-2015, 05:48 PM
Replies: 2
Views: 2,056
Posted By kenietz
strand-specific RNA-seq, tophat, RSeQC

Hi guys,
i know there are similar threads about that problem and yet it seems that is still quite confusing. Hell, is still confusing to me so i'd like to share my experience with it.

So i got...
Forum: Bioinformatics 02-26-2014, 06:59 PM
Replies: 2
Views: 2,571
Posted By kenietz
Hi guys, i am sorry to revive this thread after...

Hi guys,
i am sorry to revive this thread after long time but i also encountered this problem while trying to generate histograms on merged root file with CNVnator_0.3. Wrote to the author and...
Forum: RNA Sequencing 06-06-2013, 05:14 PM
Replies: 38
Views: 18,499
Posted By kenietz
Hi YazBraimah, glad to hear that it worked for...

Hi YazBraimah,
glad to hear that it worked for you :)

Cheers
Forum: Epigenetics 06-06-2013, 12:49 AM
Replies: 6
Views: 4,548
Posted By kenietz
I suppose it depends on the project. In some...

I suppose it depends on the project. In some cases its better to construct a special version of the genome containing only the targeted regions.

In my case of amplicon seq, 50% of the reads are...
Forum: Bioinformatics 05-13-2013, 05:12 PM
Replies: 26
Views: 6,628
Posted By kenietz
There is also the IDBA assembler: ...

There is also the IDBA assembler:

http://i.cs.hku.hk/~alse/hkubrg/projects/idba_ud/

Seems to work pretty well and does not use a lot of memory. I used it for denovo RNA-seq. Good sized...
Forum: RNA Sequencing 03-25-2013, 08:15 PM
Replies: 38
Views: 18,499
Posted By kenietz
Hi Lizex, im sorry for the late reply, im very...

Hi Lizex,
im sorry for the late reply, im very busy lately and could not find time. But better later than never :)

firstly it seems to me you compiled samtools before, i mean when you first...
Forum: RNA Sequencing 01-14-2013, 05:53 PM
Replies: 2
Views: 2,643
Posted By kenietz
Hi, i am sorry to revive that thread but i got...

Hi,
i am sorry to revive that thread but i got the same weird results. In one sample i get FPKM for a transcript and in the other not even tho when i check with IGV i get the same picture as the...
Forum: Bioinformatics 01-02-2013, 06:49 PM
Replies: 7
Views: 6,013
Posted By kenietz
Hi, firstly i'm sorry for reviving the thread...

Hi,
firstly i'm sorry for reviving the thread but i have a more specific question on the subject.

Further i will use the notation used by TOMC in previous post.

So i blast query a against set...
Forum: RNA Sequencing 11-08-2012, 05:08 PM
Replies: 38
Views: 18,499
Posted By kenietz
Hi, well i created a script which extracts a...

Hi,
well i created a script which extracts a consensus seq. I think it works but you may try and see if it works for you. Comments and suggestions are welcomed.

I attach here the script.

But...
Forum: Bioinformatics 10-11-2012, 08:26 PM
Replies: 1
Views: 1,024
Posted By kenietz
Question cufflinks: reports non existing transcripts

Hi guys,
i just discovered a very strange thing.

Im using TopHat v2.0.5, cufflinks v2.0.2, samtools-0.1.18. Ref genome and GTF from Ensembl.

I have some PE RNA-seq so i mapped it with tophat...
Forum: RNA Sequencing 10-10-2012, 12:37 AM
Replies: 47
Views: 45,993
Posted By kenietz
I see. The problem is that i cant pool them...

I see. The problem is that i cant pool them together and assemble. Too large sets. One of them is like 75M 2x75 PE and even soap-denovotrans seg faulted after loading more than 100M reads. So i had...
Forum: Bioinformatics 10-10-2012, 12:15 AM
Replies: 7
Views: 2,719
Posted By kenietz
Hi, that is what is bugging my brain as well...

Hi,
that is what is bugging my brain as well these days :D

Maybe the math is not working properly with that zero value. But thats a guess only. Came to the idea to replace all the zeros with...
Forum: RNA Sequencing 10-10-2012, 12:01 AM
Replies: 47
Views: 45,993
Posted By kenietz
Hi dvanic, thank you for the info. Well in...

Hi dvanic,
thank you for the info.

Well in fact the 3 samples were sequenced 2 times because of some trouble with the machine. The first run seems to be better tho. So i can say that i got...
Forum: RNA Sequencing 10-09-2012, 09:36 PM
Replies: 47
Views: 45,993
Posted By kenietz
Hi guys, this thread was really informative to...

Hi guys,
this thread was really informative to me. I have one question tho. I have PE rna-seq from a species without ref genome and annotation. I have only 3 samples coming from different conditions...
Forum: Bioinformatics 10-04-2012, 06:42 PM
Replies: 5
Views: 2,435
Posted By kenietz
Ok, after a lot of digging around. I reached some...

Ok, after a lot of digging around. I reached some conclusions.

1. Samtools mpileup skips reads which are marked as:
0x0100 s the alignment is not primary
0x0200 f the read fails platform/vendor...
Forum: Bioinformatics 10-01-2012, 11:33 PM
Replies: 5
Views: 2,435
Posted By kenietz
Hi rboettcher, thanks, i was reading the manual...

Hi rboettcher,
thanks, i was reading the manual for pileup as well but in my version there was no such an explanation: http://samtools.sourceforge.net/pileup.shtml

Btw, i tried using the -r but...
Forum: Bioinformatics 10-01-2012, 09:21 PM
Replies: 5
Views: 2,435
Posted By kenietz
Question samtools, cufflinks, transcripts extraction from bam

Hi guys,
i am trying to create a script which can extract the transcripts assembled by cufflinks using perl and samtools.

I tried several approaches:

1. samtools mpileup - using the pipeline...
Forum: RNA Sequencing 10-01-2012, 12:17 AM
Replies: 38
Views: 18,499
Posted By kenietz
Hi, just figured out that my work around does...

Hi,
just figured out that my work around does not work for paired-end reads :(
Somehow pileup -c removes some reads except these ones:

0x0100 s the alignment is not primary
0x0200 f the read...
Forum: RNA Sequencing 09-28-2012, 01:03 AM
Replies: 38
Views: 18,499
Posted By kenietz
Hi figo1019, here it is. Hope it does the job...

Hi figo1019,
here it is. Hope it does the job for you.

I've put some explanations inside the script. Please take a look before using it.

Comments and questions are also welcomed :)

Cheers
Forum: Bioinformatics 09-26-2012, 08:16 PM
Replies: 4
Views: 1,934
Posted By kenietz
Ok now i got it: so genes 681 and 682 have the...

Ok now i got it:
so genes 681 and 682 have the same reads mapped onto them. So when the read has FLAG field = 272 which is 100010000, is filtered out cos is nor primary alignment(bit number 9).
...
Forum: Bioinformatics 09-26-2012, 06:38 PM
Replies: 4
Views: 1,934
Posted By kenietz
Neither did i find any quality filter in...

Neither did i find any quality filter in cufflinks.
The -Q option is: min base quality (possibly capped by BAQ) [13], but it seems that is not that the problem. In IGV i see for gene682 which is...
Forum: Bioinformatics 09-26-2012, 01:26 AM
Replies: 4
Views: 1,934
Posted By kenietz
Question consensus seq, samtools pileup, transcripts.gtf issues

Hi guys,
i have some issues with extracting consensus seq for transcripts from cufflinks from the bam file.

Progs i used:
samtools-0.1.16
tophat-2.0.5
cufflinks-2.0.2
samtools-0.1.18
...
Forum: RNA Sequencing 09-16-2012, 07:29 PM
Replies: 0
Views: 1,153
Posted By kenietz
cufflinks, gff and bam order

Hi guys,
i have a simple question regarding cufflinks, bam file and gff. Its all connected to the FPKM 0 values for some transcripts but which have reads when inspect the region in the browser.
I...
Forum: RNA Sequencing 09-12-2012, 11:27 PM
Replies: 38
Views: 18,499
Posted By kenietz
Hi guys, this thread was really useful for...

Hi guys,
this thread was really useful for creating my script for extracting consensus sequences from the bam file. I tried to use the new samtools mpileup but it didnt do the job that i wanted. I...
Forum: RNA Sequencing 08-30-2012, 01:38 AM
Replies: 0
Views: 991
Posted By kenietz
Question cufflinks, transcripts extraction

Hi guys,
some clients want me to extract the consensus sequence of the transcripts assembled with cufflinks. Well i've searched around and found a way how to extract the consensus using mpileup. But...
Showing results 1 to 25 of 85

 


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