Chris:LabNotes/Moleculo/2013/2013-5-9

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Quick and Dirty Assembly Oncomelania Illumina contigs + moleculo data[edit]

Background[edit]

  • Dr. Zhang previously performed soapdenovo assembly on a smaller Illumina dataset, found in </media/LTS_15T/Kun_LTS/Snail/HB_species/Snail.HB.Soap.trimmed.K27.scafSeq>
  • We wanted to quickly check how incorporating moleculo data will help improve assembly, so we used celera assembler to do OLC assembly on contigs>100bp and all of the moleculo data (without filtering, using default settings)
  • Test run is found in my genemapster: </media/disk-2/cjwei/130419_Oncomelania_Illumina_Moleculo/Illumina_contigs+Moleculo_Assembly>

Procedure[edit]

  • Convert Illumina contigs to fastq file format using converter utility (will create artificial fastq by assigning quality score of I to each base) -> this is required for inputting file into Celera
    • Go into folder <~/software/Hybrid_error_correction_assembly_software/wgs_download_2>
    • Use command:
java convertFastaAndQualToFastq input_file > output_file
  • Combined both Illumina contigs fastq file + Moleculo fastq file into new file: <Illumina_contig+Moleculo.fastq> (using cat command)
    • It is necessary to have the correct ".fastq" file extension in order to continue forward
  • Create .frg file for input to celera (this is the primary input for assembly that gives general data on the read file)
/media/2TB_store1/cjwei/software/Hybrid_error_correction_assembly_software/wgs_download_2/wgs-7.0/Linux-amd64/bin/fastqToCA -libraryname illumina_contigs_moleculo -reads Illumina_contig+Moleculo.fastq >Illumina_contig+Moleculo.frg
    • Used "-reads" option because mimic single end reads for assembly
  • RunCA to do assembly on combined dataset
~/software/Hybrid_error_correction_assembly_software/wgs-7.0/Linux-amd64/bin/runCA -d Illumina_contig+Moleculo_assembly -p Illumina_contig+Moleculo Illumina_contig+Moleculo.frg
    • Without .spec file input, we are not doing correction as in normal hybrid assembly with pacbio+illumina data

Results[edit]

assembly n n:100 n:N50 min median mean N50 max sum
Illumina contig + Moleculo 10322 10322 2812 1000 2458 3094 3682 17558 31.94e6
Illumina contig 2167043 2167043 785219 100 132 152 148 4661 329.4e6
  • Results show a brief picture of the result of combining moleculo data with Illumina contigs. Overall, there is a dramatic increase in N50 and max contig length. However, there is a decrease in total bp assembled, probably because smaller contigs are being thrown out.
  • Will need to run more detailed tests to better assemble the data. Brief future plan:
    • Optimize assembly of Illumina reads using complete dataset
    • Run SEQuel on the assembled contigs to improve assembly accuracy
    • Hybrid assembly of Illumina contigs + Moleculo data -> will need to optimize assembly parameters for celera or explore other assemblers
    • Assembly analysis
  • Will formulate more detailed plan and discuss with Dr. Zhang