Corona Lite Introduction...General – Global SETS Versus Corona Lite Category SOLiD Global SETS v3.0 Corona_Lite v4.2 Supported OS Linux CentOS, Scyld Clusterware, PBS (Torque) Will

Corona Corona LiteLite IntroductionIntroduction

2 © 2009 Applied Biosystems

Section Outline Section Outline –– Corona Corona LiteLite IntroductionIntroduction

• Workflow and Setup

• Matching pipeline

• Pairing pipeline

• Variation pipeline


Corona Corona LiteLite OverviewOverview


GlobalSETSGlobalSETS versus Corona versus Corona LiteLite

Category SOLiD™ Global SETS v3.0 Corona_Lite v4.2

Mapping Algorithm MapReads MapReads

Mapping scheme -

progressive

Yes, for max throughput. (default)

No

-full length with fixed number of mismatches

Yes Yes

Repeat Classifier Yes, new in v3.0 No

MatchingRepeat, Random,

and Consolidate

Yes, new in v3.0 No

SNP algorithm diBayes SNP caller

Multiple run combination

analysis

No Yes.

Integrated small indel

analysis

Jun-09 Yes


Global SETS Versus Corona Global SETS Versus Corona LiteLite

Category SOLiD™ Global SETS v3.0 Corona_Lite v4.2

Matching

- Fasta-like .ma files- gff v.2

YesDefault

YesOptional using MaToGff.sh

Pairing

- .mates - gff v.2

YesDefault

YesOptional using MatesToGff.sh

SNP pipe

- SNP summary- Consensus base

sequence

Gff v.3Yes

SNP list text fileYes

Stats Files New format Old Stats file


• Before you start

• Set the correct environment

• Make cmap file

• Validate reference

• Generate double encode reference

Corona Corona LiteLite SetupSetup


• Set up environment variables

• for csh/tcsh:

• setenv CORONAROOT /share/apps/corona_lite

• source $CORONAROOT/etc/profile.d/corona.csh

• for sh/ksh/bash:

• export CORONAROOT=/share/apps/corona_lite

• source $CORONAROOT/etc/profile.d/corona.sh

Corona Corona LiteLite Setup Setup –– Environment VariablesEnvironment Variables


1 chr1 /path/to/file/chr1.fa /path/to/file/de_chr1.fa2 chr2 /path/to/file/chr2.fa /path/to/file/de_chr2.fa3 chr3 /path/to/file/chr3.fa /path/to/file/de_chr3.fa4 chr4 /path/to/file/chr4.fa /path/to/file/de_chr4.fa

• Prepare the chromosome map file (tab-delimited):

• Chromosome ID

• Chromosome Name

• FASTA Reference

• Double-Encoded Reference

• For example

Corona Corona LiteLite Setup Setup –– Chromosome Map (Chromosome Map (cmapcmap) File) File


• Validate reference

• reference_validation.pl –r chr1.fa –s 9999999999 –o chr1_validated.fa

• Generate double-encoded sequence

• encodeFasta.py -n -l sequence.fasta > de_sequence.fasta

Corona Corona LiteLite Setup Setup –– Validate and Double Encode RefValidate and Double Encode Ref








Things To ConsiderThings To Consider

• Number of hits to report (-z)

• Default is 10 per chromosome

• What does it mean if it hit 10 times?

• Recommended mismatches

• 2 for 25bp reads

• 3-4 for 35bp reads

• 4-6 for 50bp reads

• Can consider counting valid adjacent mismatches as 1 (-a=1)


Matching Parameters (Required)Matching Parameters (Required)

• matching_large_genomes_cmap_save.pl

• -csfasta – F3 or R3 reads

• -dir – Output directory

• -cmap – Chromosome map file

• -t – Tag length

• -e – Number of errors allowed


• matching_large_genomes_cmap_save.pl

• -p – Pattern mask for reads

• -a – 0 = no; 1 = valid adjacent errors; 2 = all adjacent errors: defaults to 0

• -z – Maximum number of hits per chromosome: defaults to 10

• -incremental – Remove reads that have already mapped

Matching Parameters (Optional)Matching Parameters (Optional)


Submitting JobsSubmitting Jobs

• For PBS, use submit_scripts_to_PBS.pl

• Submission scripts exist for LSF, SGE and SMP machines

• Required Options

• -j – Job list file

• Optional Options

• -h – Usage description

• -q – Specify a queue

• -i – Interactive queue








• pairing_by_group.pl

• -F3 – F3 match file (.csfasta.ma)

• -R3 – R3 match file (.csfasta.ma)

• -e – Total errors allowed during mapping

• -output_dir – Output directory

• -find_pairing_dist – Flag for finding distance distribution

• Look at pairingDist.freq.binned file

Find Insert SizeFind Insert Size


Insert Size DistributionInsert Size Distribution

0

100

200

300

400

500

600

700

800

0 500 1000 1500 2000 2500 3000


• pairing_by_group.pl

• -F3 – F3 match file (.csfasta.ma)

• -R3 – R3 match file (.csfasta.ma)

• -e – Total errors allowed during mapping

• -output_dir – Output directory

• -min_insert_size – From distribution

• -max_insert_size – From distribution

• -ref – Multi FASTA reference file

Perform MatePerform Mate--pair Rescuepair Rescue


MateMate--pair Descriptionspair Descriptions

• Mate-pairs are annotated with a three letter code








• Preparation

• Single tag: split_by_chromosome.pl

• -f – Unique match file (.unique.csfasta.ma)

• -c – Output chromosome directory

• Mate pair: multi_chr_pairing_parser.pl

• -mates – Mates file from pairing pipeline (.mates)

• -o_dir – Output directory

SNP PipelineSNP Pipeline


• Consensus and SNP calling

• consensus_prep_and_wrapper_cmap_save_script.pl

• -mates/match_dir – Output from preparation step

• -cmap – Chromosome map file

• -mlf3/mlr3 – Tag length

• -ef3/er3 – Mismatches allowed

• -o_dir – Output directory

• -insert_start/_end – Pairing size for mate pair run




• Consensus sequence generated from alignment to the reference sequence

• Files

• snps.txt

• snps_sorted.txt

• snp_probs.dat

• bp_consensus_confirmed_sequence_with_Ns.fasta


Corona Corona LiteLite OverviewOverview


QuizQuiz

• What do you need to do before running Corona Lite?

• What are the three main steps of Corona Lite?

• What is the workflow of each pipeline in Corona Lite?

• What is the meaning of the three letter annotations of mate-pairs (e.g., AAA, ABA, etc)?

• What are the main differences between Corona Lite and GlobalSETS?

AppendixAppendix


General General –– Global SETS Versus Corona Global SETS Versus Corona LiteLiteCategory SOLiD™ Global SETS v3.0 Corona_Lite v4.2

Supported OS Linux CentOS, ScyldClusterware, PBS (Torque)Will test LSF, PBS pro and SGE by June 2009

Linux, PBS, LSF, SGE

Programming language Java (algorithms in C++) Scripting languages (some algorithms in C++)

Analysis set up and

execution

Automatic through SETS GUI;Integrated command line

Integrated command. Can run batch mode.

Integrate with custom pipeline

Yes (SAI)GUI, and Command line

Yes, command line interface

Speed Optimized for compute performance for complex genome analysis

Support complex genome analysis

Warranty Yes No

AB support to end users Yes Yes

License Fee Comes with SOLiD 3 System Contact AB sales

Free open source

Documents

Corona Lite Introduction...General – Global SETS Versus Corona Lite Category SOLiD Global SETS v3.0 Corona_Lite v4.2 Supported OS Linux CentOS, Scyld Clusterware, PBS (Torque) Will