output for sequencing of reverse strand

[bio_informatics]

Hi Members,

I've lost, yet again with direction, and sequencing. I'm working on 16s data. Paired end, Illumina reads. And since then I am confused over the output in read 2- fastq.gz

5'------>primer SEQUENCE END 3'
3' END SEQUENCE PRIMER<-------5'

Paired end sequencing as per my understanding looks like above. Primer or index, or bar code, adapter, Pardon me for not using appropriate terms.

My doubt:
1) In the fastq of reverse strand, how do I have the output:
END.............SEQUENCE
OR
SEQUENCE......END

I'm confused over the direction in which I have the fastq output.

[GenoMax]

DNA strands are anti-parallel. So R2 in your cartoon has the 5'-end to the right of that strand (and not 3' as you have drawn it).

See this for a simple explanation: http://seqanswers.com/forums/showthread.php?t=42737

[bio_informatics]

Hi Genomax,

Thanks for your reply and sharing the URL.
I've edited my cartoon. Sorry.

But then again, in which direction is the output present for reverse read in fastq?

[GenoMax]

DNA sequence is always represented as 5' --> 3'. So 5'--SEQUENCE ... END--3' for R2.

[bio_informatics]

DNA sequence is always represented as 5' --> 3'. So 5'--SEQUENCE ... END--3' for R2.

Thanks for clearing clouds.

That explains, why reverse read file has bar code/index at the starting of reads.

Merci.!

[GenoMax]

Thanks for clearing clouds.

That explains, why reverse read file has bar code/index at the starting of reads.

Merci.!

You must be using a custom "inline" barcode/index. Illumina barcodes are never part of actual read.

[bio_informatics]

You must be using a custom "inline" barcode/index. Illumina barcodes are never part of actual read.

Hi Genomax,

Yes, I guess so. Thanks for sharing new term. Everyday a new learning.

Data am having is based on entirely new chemistry/protocol. Here's the paper.