Short read toolbox Botany2010

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This is a legacy website!

This page was originally created to provide resources for a presentation at the the Botany 2010 conference. It has been moved here to preserve it for those users who are still interested in accessing it. This is now a legacy website. For current information please see:

Short read toolbox

Short Read Toolbox

This page has been created to help list resources for working with high throughput sequencing data. You can also check out our individual lab pages to see updates on methods -- Cronn Lab or Liston:Lab -- to see updates on methods .

Short Read Workshop

Download Presentations and Training Modules from our Recent Short Read Workshop, "An introduction to next-generation sequencing". Presented at the Botany 2010 Meeting in Providence, R.I.


Currently available platforms:

  • Illumina - Illumina (formerly Solexa).
  • 454 - 454/Roche.
  • SOLiD - ABI by Life Technologies.

Anticipated technologies:

Online short-read resources

Sequence format information

  • Short Read Toolbox - Descriptions and examples of qseq, scarf, fastq and fasta formats. Includes scripts to translate these formats to the fastq format standard.
  • FASTQ - Wikipedia's FASTQ page.
  • FASTA - Wikipedia's FASTA page.

Alignment format information

Short-read quality control software

  • TileQC - Requires R, RMySQL and MySQL.
  • FastQC - A quality control tool for high throughput sequence data. A Java application.
  • Short Read Toolbox - Scripts for quality control of Illumina data.

Open source de novo assemblers

  • Velvet - Implements De Bruijn Graphs in C. Requires 64 bit Linux OS.
  • Edena - 32 and 64 bit Linux.
  • ABySS - Multi-threaded de novo assembly.
  • Ray - Multi-threaded de novo assembly.
  • QSRA - Utilizes quality scores.

Open source reference guided assemblers

  • SOAP - Short Oligonucleotide Analysis Package.
  • MAQ - Mapping and Assembly with Qualities.
  • Bowtie - Bowtie. An ultrafast, memory-efficient short read aligner.
  • BWA - Burrows-Wheeler aligner.
  • RGA - Perl script which calls blat to assemble short reads.

Hybrid assemblers (reference guided & de novo)

RNA-Seq / Transcriptome

  • TopHat - A fast splice junction mapper for RNA-Seq reads.
  • Cufflinks - Assembles transcripts, estimates their abundances, and tests for differential expression and regulation.
  • SuperSplat - Splice junction discovery.

Assembly viewers

  • Tablet - Tablet, visualizes ACE, AFG, MAQ, SOAP, SAM and BAM formats.
  • SAMtools - SAMtools.

Alignment programs

  • T-Coffee - T-Coffee.
  • Muscle - Muscle.
  • LASTZ - LASTZ, hosted at the Miller lab.
  • MUMmer - MUMmer.
  • Mulan Multiple Sequence Alignment and Visualization Tool.
  • VISTA Tools for Comparative Genomics.
  • mauve - Multiple (bacterial) genome aligment.

Sequence query programs

  • PLAN - A web application for conducting, organizing, and mining large-scale BLAST searches (limited to 1,000 queries).
  • BLAT - BLAT.



A very brief example to demonstrate file input/output.


use strict;
use warnings;
my (@temp, $in, $out);
my $inf = "data.fq";
my $outf = "data_out.fq";
open($in, "<", $inf) or die "Can't open $inf: $!";
open($out, ">", $outf) or die "Can't open $outf: $!";
  chomp($temp[0]=$_); # First line is an identifier.
  chomp($temp[1]=<$in>); # Second line is sequence.
  chomp($temp[2]=<$in>); # Third line is an identifier.
  chomp($temp[3]=<$in>); # Fourth line is quality.
  print $out join("\t", @temp)."\n";
close $in or die "$in: $!";
close $out or die "$out: $!";
  • perlintro - Introduction to perl with links to other documentation.
  • BioPerl beginners - Introduction to BioPerl (be prepared for object oriented code).


R project

Useful links