######################################################### # README - # Reference-guided de novo assembly # ==================================================== # by Heidi Lischer (heidi.lischer@ieu.uzh.ch), 2015/15 ######################################################### We adapted and extended the reference-guided assembly approach from Schneeberger et al. [1]. The main idea of this approach is to first map reads against a reference genome of a related species to reduce the complexity of de novo assembly within continuous covered regions. In a further step, reads with no similarity to the related genome are integrated. Reference-guided de novo assembly pipeline: 1. Step: quality/adapter trimming and quality check 2. Step: map reads against reference and define blocks and superblocks 3. Step: do deNovo assembly within superblocks 4. Step: get non-redundant supercontigs 5. Step: map reads on supercontigs and de novo assemble unmapped reads 6. Step: map reads to all supercontics and correct them 7. Step: scaffolding and gap closing [1] http://www.pnas.org/content/early/2011/06/01/1107739108.full.pdf+html?with-ds=yes Dependencies -------------- (in brackets: used version of softwares) Third party programs: - fastqc (v 0.10.1): Fastq quality checking - trimmomatic-0.32 (v 0.32): quality and adapter trimming - samtools (v 1.3): Tools for alignments in the SAM format - bcftools (v 1.3): Tools for variant calling and manipulating VCFs and BCFs - bamtools (v 2.3.0): Tools for alignments in the BAM format - bedtools (v 2.19.0): Tool for genomic arithmetics - picardtools (v 1.109): Processing alignment files - bowtie2 (v 2.2.1): NGS alignment tool - seqtk (v 1.0-r45): Fast Fasta/Fastq manipulation tool - AMOScmp (v 3.1.0): Comprarative genome assembly - MUMmer (v 3.23): Comprarative genome assembly - GenomeAnalysisTK (v 3.1-1): Genome analysis toolkit - SOAPdenovo2 (v r240): De novo genome assembler and scaffolder and one of these de novo assemblers: - AllPaths-LG (v 51279) - idba (v 1.1.1) - abyss-pe (v 1.5.2) - SOAPdenovo2 (v r240) Additional in-house scripts ----------------------------- - RemoveShortSeq.jar: Remove short sequences and make unique identifiers from FASTA/FASTQ - GetBlocks_new.jar: Create blocks and superblocks - FastaToAmos.jar: Transform FASTA to Amos format - WriteSoapConfig.jar: writes a SOAP config file - FastaStats.jar: outputs FASTA statistics - SplitSeqLowCov.jar: splits sequences at low coverage Running --------- In first lines of the main scripts you have to adapt the paths to your system. (Everything between: # set variables ######################################### ... #########################################################) If the parameters are set you can run the pipeline as follows: bash refGuidedDeNovoAssembly_ALLPATHS.sh or bash refGuidedDeNovoAssembly_IDBA.sh or bash refGuidedDeNovoAssembly_ABYSS.sh or bash refGuidedDeNovoAssembly_SOAP.sh