RNA-seq Quality Control help

[lewewoo]

I am going to see the quality of the RNA-seq data, is any method or program available to help me to do this? Thanks!

I heard some people analyze the distribution of ATCG in the different position of the reads... or compare the lane's complexity...

[lewewoo]

ShortRead could assess the data quality; and other stuffs could be used? How to evaluate the ribosomal RNA contaminations.... Thanks!

[ttnguyen]

I often use fastQC for this task.

http://www.bioinformatics.bbsrc.ac.uk/projects/fastqc

[ttnguyen]

But I am not sure if fastQC can help with the ribosomal RNA contaminations.... I think there are some discussions about that in this forum.

[lewewoo]

I will start from FastQC; any more packages for QC control? Thanks a lot!

[lewewoo]

interpreting the QC

Thanks again for your help!
My samples were done in Illumina and I have used FastQC to check the quality of my samples, I have some questions as for 1) per sequence quality scores; 2)Sequence Duplication Levels;

Q1: there are up-down peaks starting from position 1 to position 15, and my sequence length is 75nt; is this OK? I heard people said before 16 there are always some nosies....

Q2: Sequence duplication levels and Kmer Content? first one is big than 69 and second one has some problems with the beginning sequences; is this also related to Q1


I really appreciate all the helps from this forum, thanks a lot!

I often use fastQC for this task.

http://www.bioinformatics.bbsrc.ac.uk/projects/fastqc

[Puva]

Regarding rRNA contamination-
I am not sure whether there is a database for ribosomal RNA like other RNA species(miRNA and ncRNA). If available, data can be blast against this database to see the contamination.

[liguow]

EVER-seq (Evaluate Experiment of RNA-seq)

I often use fastQC for this task.

http://www.bioinformatics.bbsrc.ac.uk/projects/fastqc

fastqc only check the quality of sequence per se (such as GC content, base calling quality, over represented kmer, etc), that's fundamental and important. However, fastqc does not tell you if the RNA-seq experiment was successful or not (it only tells you whether the sequencing was successful).

We develop a package to QC RNA-seq experiment from different aspects such as: gene body coverage, reads distribution over the genome, duplication rate, reproducibility and saturation etc.

http://code.google.com/p/ever-seq/

We change the name of EVER-seq into RSeQC. Because Google Code doesn't allow rename an exist project, we create a new project with the url:

http://code.google.com/p/rseqc/

From now on, we will update and add new modules to RSeQC

[Jon_Keats]

The picard tool set also has a nice RNAseq metrics tool

[lukas1848]

You could also use the perl script preprocessest.pl with the -dr option set for the removal of ribosomal cDNA. You'd need to set up a blast db with your organism's rDNA first though.