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README.txt

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+Cerulean Hybrid Genome Assembler v0.1.1
+
+This software extends contigs assembled using short read datasets like Illumina
+paired-end reads using long reads like PacBio RS long reads.
+
+The method is fully described in:
+Deshpande, V., Fung, E. D., Pham, S., & Bafna, V. (2013).
+Cerulean: A hybrid assembly using high throughput short and long reads.
+arXiv preprint arXiv:1307.7933.
+
+A] Requirements:
+Ubuntu 12.04 (may run on other operating systems, but not tested)
+Python 2.7.1 (may run on older versions, but not tested)
+numpy, matplotlib libraries for Python
+ABySS assembler: http://www.bcgsc.ca/platform/bioinfo/software/abyss
+SMRT Analysis tookit (for BLASR): http://pacbiodevnet.com/
+PBJelly: https://sourceforge.net/projects/pb-jelly/ 
+
+B] Inputs and Pre-processing:
+ i) Assembled contigs from ABySS short read assembler
+ ii)Mapping of Pacbio reads to ABySS contigs using BLASR
+
+ i) Assembly of Illumina paired-end reads:
+   If the paired-end reads are stored in fastq format in the files reads1.fastq
+   and reads2.fastq, then contigs may be assembled by:
+   $ abyss-pe k=64 n=10 in='reads1.fastq reads2.fastq' name=<dataname>
+   This will generate 2 files used for inputs to Cerulean:
+   * <dataname>-contigs.fa    #This contains the contig sequences
+   * <dataname>-contigs.dot   #This contains the graph structure
+
+ ii)Mapping PacBio reads to ABySS contigs using BLASR:
+   Note: sawriter and blasr are part of SMRT Analysis toolkit
+   Note: You need to set the environmental variables and path:
+   $ export SEYMOUR_HOME=/opt/smrtanalysis/
+   $ source $SEYMOUR_HOME/etc/setup.sh
+
+   Suppose PacBio reads are stored in <dataname>_pacbio.fasta
+   $ sawriter <dataname>-contigs.fa
+   $ blasr <dataname>_pacbio.fa <dataname>-contigs.fa -minMatch 10 \
+     -minPctIdentity 70 -bestn 30 -nCandidates 30 -maxScore -500 \
+     -nproc <numthreads> -noSplitSubreads \
+     -out <dataname>_pacbio_contigs_mapping.fasta.m4
+
+   Make sure the fasta.m4 file generated has the following format:
+   qname tname qstrand tstrand score pctsimilarity tstart tend tlength \
+   qstart qend qlength ncells
+   The file format may be verified by adding the option -header to blasr. 
+
+C] Execute:
+ Cerulean requires that all input files are in the same directory <basedir>:
+ i)   <basedir>/<dataname>-contigs.fa
+ ii)  <basedir>/<dataname>-contigs.dot
+ iii) <basedir>/<dataname>_pacbio_contigs_mapping.fasta.m4
+
+ To run:
+ $ python src/Cerulean.py --dataname <dataname> --basedir <basedir> \
+ --nproc <numthreads>
+
+ This will generate:
+ i)  <basedir>_cerulean.fasta
+ ii) <basedir>_cerulean.dot
+ Note: The dot does not have same contigs as fasta, but intermediate graph.
+
+
+D] Post-processing:
+ Currently Cerulean does not include consensus sequence of PacBio reads in gaps
+ The gaps may be filled using PBJelly.
+ $ python $JELLYPATH/fakeQuals.py <dataname>_cerulean.fasta <dataname>_cerulean.qual
+ $ python $JELLYPATH/fakeQuals.py <dataname>_pacbio.fasta <dataname>_pacbio.qual
+ $ cp $JELLYPATH/lambdaExample/Protocol.xml .
+ $ mkdir PBJelly
+ Modify Protocol.xml as follows:
+ Set <reference> to $PATH_TO_<basedir>/<dataname>_cerulean.fasta 
+ Set <outputDir> to $PATH_TO_<basedir>/PBJelly
+ Set <baseDir> to $PATH_TO_<basedir>
+ Set <job> to <dataname>_pacbio.fasta
+ Set <blasr> option -nproc <numthreads> 
+ Note: PBJelly requires that the suffix be .fasta and not .fa
+ Next run PBJelly:
+ ($ source $JELLYPATH/exportPaths.sh)
+ $ python $JELLYPATH/Jelly.py <stage> Protocol.xml
+ where <stage> has to be in the order:
+ setup
+ mapping
+ support
+ extraction
+ assembly
+ output
+
+ The assembled contigs may be view in <basedir>/PBJelly/assembly/jellyOutput.fasta
+
+In case of any questions or errors please contact vdeshpan eng DT ucsd DT edu