Go Back   SEQanswers > Bioinformatics > Bioinformatics

Similar Threads
Thread Thread Starter Forum Replies Last Post
bad quality reads in reverse direction using SureSelect on MiSeq somm Introductions 1 11-03-2016 03:58 AM
[question] reads detected sequences out of sequencing primer in the other direction SF_mallish Bioinformatics 2 12-06-2012 01:55 PM
How to remove reads in wrong direction from Cufflinks analysis Noa Bioinformatics 0 03-01-2012 01:21 AM
SOLiD SAGE reads direction and orientation invermay SOLiD 0 02-14-2011 06:17 PM
Single direction reads for variants Moggs General 11 06-22-2010 06:55 AM

Thread Tools
Old 08-16-2017, 10:39 AM   #1
Senior Member
Location: US

Join Date: Apr 2013
Posts: 222
Default Should I worry about the direction of my reads

Hello, I recently got my metagenomic illumina sequencing back (hiseq). There is one sample that I would like to do some blast. Since it was sequenced using paired-end sequencing (I have two fastq files forward and reverse sequencing) for this sample, here is what I am going to do.

First, I interleaved the two fastq files using bbmap. So, I will get one fastq file and easy for me. Next, I plan to use the file to do blast.

Here is my concerned. When I run BBmap, I didn't notice if there is no option to sort the direction of the two fastq files. For example, I want to make all reads in the interleaved fastq file have the same direction (e.g., 5'-3' or 3'-5'). I am concerned if this interleaved fastq file mixed with reads having different directions could make the NCBI blast program failed to find the best match in the database.

Here are my questions:

1>The reads in the two fastq files (forward and reverse) that I got from the sequencing center are same directions or opposite directions? I know the machine sequenced from opposite direction. How about the reads directions in the fastq files? Already sorted? How can you tell? Are they from 3' to 5' or 5' to 3'?

2>Are the common software (e.g., BLAST+, RApsearch, DIANMOND) consider the direction problem, when they do search?

SDPA_Pet is offline   Reply With Quote
Old 08-17-2017, 05:35 AM   #2
Senior Member
Location: USA, Midwest

Join Date: May 2008
Posts: 1,178

Unless it is explicitly stated that a reported DNA sequence is presented in a 3' to 5' orientation it is accepted convention that all DNA sequences are written 5' to 3'. All sequence data reported by an Illumina (or other) sequencer is in 5'->3' orientation. BLAST or any other sequence alignment program expects and assumes that your input FastQ files are 5'->3'. These programs all internally manage the alignment of the query to the reference in both the its input orientation and its reverse complement.
kmcarr is offline   Reply With Quote

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off

All times are GMT -8. The time now is 04:36 AM.

Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2021, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO