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Forum: Bioinformatics 07-02-2015, 07:36 AM
Replies: 1
Views: 2,088
Posted By PFS
Find mates in a SAM file

I want to identify the actual pairs of reads in a SAM file that potentially has multiple alignments for the same given pair.

Is it sufficient to check whether two lines in the SAM file have same...
Forum: Bioinformatics 02-24-2012, 05:25 PM
Replies: 10
Views: 5,895
Posted By PFS
skipping cufflinks-->cuffcompare ... straight to cuffdiff?

I see that most workflows includes tophat --> cufflinks --> cuffcompare --> cuffdiff.

If I want to perform differential expression analysis on RNASEQ samples based on a known annotation (e.g....
Forum: Bioinformatics 09-12-2011, 07:23 AM
Replies: 14
Views: 5,866
Posted By PFS
I thought that v.0.9 should be able to...

I thought that v.0.9 should be able to distinguish between encodings (see below) ... but I will try to see if the latest version can help.

Thanks!


From the release notes:
"30-3-11: Version...
Forum: Bioinformatics 09-12-2011, 05:52 AM
Replies: 14
Views: 5,866
Posted By PFS
FASTQC guessing wrong quality encoding

Hello,

I have some Illumina files processed with CASAVA 1.8.
The program FASTQC is guessing the format to be be Illumina 1.5
Is there a way to explicitly tell fastqc what encoding the data is?...
Forum: Bioinformatics 09-09-2011, 06:45 AM
Replies: 1
Views: 1,891
Posted By PFS
typical parameters for filtering Illumina RNASEQ reads

I've used fastq_quality_filter with q = 3 and p = 90 (at least 90% of each read must have quality >3). I thought these were relaxed settings and was expected only few reads being discarded...
Forum: Bioinformatics 09-06-2011, 08:16 AM
Replies: 4
Views: 3,849
Posted By PFS
I read more carefully the paper, and I understand...

I read more carefully the paper, and I understand that trimming the 5' end does not work.

I am curious to know whether this bias is of any real concern for differential expression analysis. If...
Forum: Bioinformatics 09-06-2011, 08:02 AM
Replies: 4
Views: 3,849
Posted By PFS
dealing with 5' end bias in RNASEQ

Hello,
I read about random hexamer priming bias in Illumina RNASEQ (see Hansen et al., NAR 2010, vol 38 no 12). Specifically, "There is a strong distinctive pattern in the nucleotide frequencies...
Forum: Bioinformatics 08-10-2011, 06:35 AM
Replies: 5
Views: 8,131
Posted By PFS
Question comparing results by cuffdiff, edgeR, DESeq

Has anyone tried to compare the results from the various tools that offers differential expression analyses for RNASEQ data?

I understand they have different underlying models and assumptions, but...
Forum: Bioinformatics 08-02-2011, 11:01 AM
Replies: 9
Views: 2,371
Posted By PFS
Thank you Andrew for the suggestion. I will try...

Thank you Andrew for the suggestion. I will try to combine the list.
Forum: Bioinformatics 08-02-2011, 10:29 AM
Replies: 9
Views: 2,371
Posted By PFS
I used edgeR to determine the list of...

I used edgeR to determine the list of up-regulated and down-regulated genes. I have 3 samples, so in total I have 6 lists. The number of genes for each list is:

388 edger_1_down.genelist
503...
Forum: Bioinformatics 08-02-2011, 08:59 AM
Replies: 9
Views: 2,371
Posted By PFS
GO enrichment RNASEQ: nothing!

Hello,

I used goseq package to do GO-enrichment of RNASEQ data. The up- and down-regulated genes are not enriched in any category if I use the BH (FDR) correction. Is that a possible situation or...
Forum: Bioinformatics 07-19-2011, 09:18 AM
Replies: 3
Views: 3,744
Posted By PFS
Bowtie call to get unique, multi-hits and nonmatching reads

Hello there,

I want to estimate (1) the number of reads that map uniquely (in one place only), (2) the number of reads that map in multiple places and (3) the number of reads that do not map at...
Forum: Bioinformatics 07-05-2011, 01:15 PM
Replies: 5
Views: 4,274
Posted By PFS
can small rnaseq data be analyzed like rnaseq data?

Sorry if this is a dumb question.

I was wondering whether small rna-seq data can be analyzed with the standard software for rna-seq data such as TopHat, Cufflinks, DESeq, etc...

Are there...
Forum: Bioinformatics 07-05-2011, 10:25 AM
Replies: 3
Views: 7,417
Posted By PFS
fastqc - overrepresented sequences

I have run a FASTQC analysis and found out that there are hundreds of over-represented sequences in my datasets. Some of these, are Illumina PCR primers, some are single end adapters.

Does it...
Forum: Bioinformatics 06-21-2011, 09:14 AM
Replies: 0
Views: 2,714
Posted By PFS
clustering RNASeq data

Hi,
I would like to do some lustering of rnaseq data.
I used tophat/cuffdiff to get a table with the fpmk values.
Beside the quantile normalization option in cuffdiff, what other trick has to be...
Forum: Bioinformatics 06-17-2011, 11:11 AM
Replies: 0
Views: 2,129
Posted By PFS
library complexity for RNASEQ

How can I establish whether my library has a good complexity? What percentage of the reads should be unique?
Thanks!
Forum: Bioinformatics 06-14-2011, 11:07 AM
Replies: 1
Views: 4,079
Posted By PFS
cuffdiff with biological replicates

The FPKM values reported by CUFFDIFF when the samples have biological replicates correspond to the average value across the replicates?

Thanks in advance
Forum: Bioinformatics 05-27-2011, 09:04 AM
Replies: 3
Views: 2,318
Posted By PFS
cuffdiff count tables

I tried the option to emit count tables in cuffdiff. I expected the counts to be integers, but I get fractions. Are these normalized counts?
Forum: Bioinformatics 05-23-2011, 12:57 PM
Replies: 4
Views: 2,115
Posted By PFS
Thanks! That's what I needed!

Thanks! That's what I needed!
Forum: Bioinformatics 05-20-2011, 01:12 PM
Replies: 4
Views: 2,115
Posted By PFS
Thank you, HESmith! The barcodes are in the...

Thank you, HESmith!

The barcodes are in the header so I am assuming I cannot use the utility in fastx_toolkit.

Does it make sense to list all possible barcodes that would be acceptable (i.e....
Forum: Bioinformatics 05-20-2011, 11:15 AM
Replies: 4
Views: 2,115
Posted By PFS
non-matching barcodes

Hi,

I have some RNASEQ data from Illumina 1.4+ pipeline.
The barcodes of a multiplex run are appended to the header line of the fastq file. I supposedly have two samples in there, so I am...
Forum: Bioinformatics 04-21-2011, 11:38 AM
Replies: 2
Views: 3,545
Posted By PFS
"coverage" of introns, intergenic regions for RNASEQ

I would like to determine the % of reads mapping to exons, introns and intergenic regions. I know BEDTools can do this, but I am not sure how to get the GFF files with the information for exons,...
Forum: Bioinformatics 04-13-2011, 06:38 AM
Replies: 4
Views: 2,504
Posted By PFS
Hi, You are right. It would make more sense...

Hi,

You are right. It would make more sense to separate TopHat from Cufflinks. I guess my intention was to pinpoint to some kind of alignment tools with or without transcriptome assembly tool.
...
Forum: Bioinformatics 04-13-2011, 05:18 AM
Replies: 0
Views: 1,151
Posted By PFS
Does the TopHat illumina quality option work?

I was never able to get the "--solexa1.3-quals" option to work. When I manually convert quality and then run TopHat everything is ok, but when I skip the manual conversion, I always get an error...
Forum: Bioinformatics 04-13-2011, 05:14 AM
Replies: 4
Views: 2,504
Posted By PFS
Which of these steps in RNA-Seq pipeline do you do?

In your analysis of RNA-Seq pipeline, which of the following steps do you normally do?

(Illumina)

PRE-PROCESSING
1) removal of reads with consecutive stretches of "B" qualities
2) trimming 3'...
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