How Can I Tell What Is The Adapter Used In A Sequence Read Archive (Sra) Sample?

Question

[Doctoroots]

Hi all, i want to perform some analysis on small RNA sequence samples retrieved from the Sequence Read Archive (SRA). since adapter contamination is common in small RNA seuquencing results, its important to clip the adapter from the retrieved fastq file before the downstream analysis. to clip the adapter, i need to know its sequence.

so my question is : how do i know what was the adapter used in the sample preparation?

an example experiment : SRAsmallRNA_experiment

[Michael]

Found by google ("illumina adapter sequence"): http://seqanswers.com/forums/showthread.php?t=198

[Doctoroots]

Hi Michael, i know this sequence list, but it seems that sometimes a different adapter is used, or a similar adapter with a few nucleotides difference at the beginning or end. i wanted to know if there is a way to find the exact adapter used. thanks.

Answer 1

[Alastair Kerr]

Run the sequences through fastQC It will also detect loads of other weirdness as well including both adaptor and primer contamination, quality problems etc. All sequences for the contaminants are present in the config file called "contaminant_list.txt"

[Michael]

Cool tool! Wish I had known about this earlier

[sckinta]

I thought the contaminant_list.txt is an input file, isn't it? There is a option on fastqc command that to specify a non-default file which contains the list of contaminants to screen overrepresented sequences against.

[Edalat]

Dear Alastaire

I use fastQC for find adapter,it reports "illumina universal adapter" for some RNA-seq and small RNA-seq data,do you know what its sequence is?

[GenoMax]

BBMap suite includes sequences of all popular adapters in a file called "adapters.fa" in resources directory you will find in BBMap folder.

[Edalat]

thank you for your attention,this file has "TruSeq_Universal_Adapter" sequence, I need sequence of "illumina universal adapter".

[Michael]

TruSeq is the name of the illumina sequencing protocol, so they are the same. Recent Hiseq and Miseq machines should be covered by TruSeq version3 adapter sequences, at least that is what works with Trimmomatic.

[Edalat]

Dear Michael

I sent an E-mail to manager of FastQC and ask him about this adapter,ha said The universal adapter isn’t actually an adapter sequence as such, it’s the common sequence which exists at the start of all of the illumina adapter and primer sequences before they diverge into the different sub-variants. This is the first sequence you encounter when you read off the end of an insert on an illumina run so you can use it for detecting and trimming adapters without having to know exactly which one was used.

it is an additional info?

or

It means I should remove 3and 5 terminal nucleotide for trim universal adapter?

[GenoMax]

Just use the sequence of universal adapter. Trimming programs will remove all sequence 3' of the point where they find the universal adapter. If you use bbbduk.sh you can point it to the entire adapters.fa file and it will do the rest.

[Edalat]

then I should use "truseq universal illumina" sequence when FastQC reports "illumina universal adapter".

I do not use software except windows software,it is better to say I use clc genomics only.

[GenoMax]

You can use that sequence with CLC according to these directions.

[Edalat]

yes,I would like to tell you that I can not use bbMap and other non-windows software.

I really appreciate your help .

[GenoMax]

BBMap is pure Java and will happily run on windows as long as you have Java installed.

[Edalat]

really?! then I should try it,however I think BBMap operate like clc for trimming adapter.