Platanus

Platanus-allee (formerly known by Platanus2)

Description

Platanus-allee is a de novo haplotype assembler (phasing tool), which assembles each haplotype sequence in a diploid genome. Compared to the read mapping-based haplotype phasing tools, Platanus-allee is especially useful to analyze highly divergent (heterozygous) regions in which haplotypes extremely differ. This tool requires at least one Illumina library and can accepts Illumina mate-pairs, PacBio/Oxford-Nanopore long reads and 10X linked-reads. In addition to the haplotype phasing function, Platanus-allee can construct consensus sequences (pseudo haploid genome), which have mosaic structures of haplotypes (i.e., maternal and paternal haplotypes are mixed) and can be used as the conventional “draft genome”. Note that Platanus-allee was previously called “Platanus2” and renamed according to emphasize the difference from Platanus, which is the tool to assemble consensus sequences of haplotypes.

Version

  • v.2.0.0

Download

  • Linux 64 bit binary (precompiled)Download

Requirements

  • Minimap2
    • https://github.com/lh3/minimap2
    • Only required to use PacBio/Oxford-Nanopore long reads.
  • Long Ranger
    • https://support.10xgenomics.com/genome-exome/software/pipelines/latest/what-is-long-ranger
    • Only required to prepare 10X linked-reads input (barcoded.fastq).

 

Synopsis
Inputs

  • Illumina paired-end: PE_1.fq PE_2.fq (mandatory)
  • Illumina mate-pair : MP_1.fq MP_2.fq (optional)
  • PacBio long reads : PacBio.fq (optional)
  • 10X linked-reads : 10X_barcoded.fq (optional)

Commands

platanus_allee assemble -f PE_1.fq PE_2.fq 2>assemble.log

 

platanus_allee phase \
-c out_contig.fa out_junctionKmer.fa \
-IP1 PE_1.fq PE_2.fq \
-OP2 MP_1.fq MP_2.fq \
-p PacBio.fq \
-x 10X_barcoded.fq \
2>phase.log

 

platanus_allee consensus \
-c out_primaryBubble.fa out_nonBubbleHomoCandidate.fa \
-IP1 PE_1.fq PE_2.fq \
-OP2 MP_1.fq MP_2.fq \
-p PacBio.fq -x 10X_barcoded.fq \
-x 10X_barcoded.fq \
2>consensus.log

Final output

out_allPhaseBlock.fa
out_consensusScaffolds.fa
out_consensusScaffoldComponent.bed

Misc. output

out_primaryBubble.fa
out_secondaryBubble.fa
out_nonBubbleHomoCandidate.fa
out_nonBubbleHetero.fa

 

Contig assembly usage
Command

platanus_allee assemble [OPTIONS] 2>log

Options

-o STR : prefix of output files (default out, length <= 200)
-f FILE1 [FILE2 …] : reads file (fasta or fastq, number <= 100)
-k INT : initial k-mer size (default 32)
-K FLOAT : maximum-k-mer factor (maximum-k = FLOAT*readlength, default 0.5)
-s INT : step size of k-mer extension (>= 1, default 20)
-n INT : initial k-mer coverage cutoff (default 0, 0 means auto)
-c INT : minimum k-mer coverage (default 2)
-a FLOAT : k-mer extension safety level (default 10.0)
-u FLOAT : maximum difference for bubble crush (identity, default 0)
-d FLOAT : maximum difference for branch cutting (coverage ratio, default 0.5)
-e FLOAT : k-mer coverage depth (k = initial k-mer size specified by -k) of homozygous region (default auto)
-t INT : number of threads (<= 100, default 1)
-m INT : memory limit for making kmer distribution (GB, >=1,default 16)
-tmp DIR : directory for temporary files (default .)

Outputs

PREFIX_contig.fa
PREFIX_junctionKmer.fa
PREFIX_kmerFrq.tsv

PREFIX is specified by -o

 

Haplotype phasing usage
Command

platanus_allee phase [OPTIONS] 2>log

Options

-o STR : prefix of output file and directory (do not use “/”, default out, length <= 200)
-c FILE1 [FILE2 …] : contig (or scaffold) file (fasta format)
-ip{INT} PAIR1 [PAIR2 …] : lib_id inward_pair_file (interleaved file, fasta or fastq)
-IP{INT} FWD1 REV1 [FWD2 REV2 …] : lib_id inward_pair_files (separate forward and reverse files, fasta or fastq)
-op{INT} PAIR1 [PAIR2 …] : lib_id outward_pair_file (interleaved, fasta or fastq)
-OP{INT} FWD1 REV1 [FWD2 REV2 …] : lib_id outward_pair_files (separate forward and reverse files, fasta or fastq)
-p PAIR1 [PAIR2 …] : long-read file (PacBio, Nanopore) (fasta or fastq)
-x PAIR1 [PAIR2 …] : linked-reads files (paired-ends, 10x Genomics) (interleaved, fasta or fastq)
-X FWD1 REV1 [FWD2 REV2 …] : linked-reads files (paired-ends, 10x Genomics) (separate forward and reverse files, fasta or fastq)
-i INT : number of iterations (default 2)
-l INT : minimum number of links to scaffold (default 3)
-k INT : minimum number of links to phase variants (default 1)
-t INT : number of threads (default 1)
-tmp DIR : directory for temporary files (default .)
-mapper FILE : path of mapper executable file (default minimap2, only effective with -p option)
-minimap2_sensitive : sensitive mode for minimap2 (default, off; only effective with -p option)

Outputs

PREFIX_allPhaseBlock.fa
PREFIX_primaryBubble.fa
PREFIX_secondaryBubble.fa
PREFIX_nonBubbleHomoCandidate.fa
PREFIX_nonBubbleHetero.fa
PREFIX_intermediateResults (directory)

PREFIX is specified by -o

 

Consensus scaffold construction usage
Command

platanus_allee consensus [OPTIONS] 2>log

Options

-o STR : prefix of output file (default out, length <= 200)
-c FILE1 [FILE2 …] : contig_file (fasta format)
-ip{INT} PAIR1 [PAIR2 …] : lib_id inward_pair_file (interleaved file, fasta or fastq)
-IP{INT} FWD1 REV1 [FWD2 REV2 …] : lib_id inward_pair_files (separate forward and reverse files, fasta or fastq)
-op{INT} PAIR1 [PAIR2 …] : lib_id outward_pair_file (interleaved, fasta or fastq)
-OP{INT} FWD1 REV1 [FWD2 REV2 …] : lib_id outward_pair_files (separate forward and reverse files, fasta or fastq)
-p PAIR1 [PAIR2 …] : long-read file (PacBio, Nanopore) (fasta or fastq)
-x PAIR1 [PAIR2 …] : linked-reads files (paired-ends, 10x Genomics) (interleaved, fasta or fastq)
-X FWD1 REV1 [FWD2 REV2 …] : linked-reads files (paired-ends, 10x Genomics) (separate forward and reverse files, fasta or fastq)
-e FLOAT : coverage depth of homozygous region (default auto)
-L INT : maximum fragment length of tag (10x Genomics) (default 200000)
-s INT1 [INT2 …] : mapping seed length for short reads (default 32 64)
-S INT1 [INT2 …] : mapping seed length for long reads (default 20, only effective with -kmer_align option)
-l INT : minimum number of links to scaffold (default 3)
-t INT : number of threads (<= 1, default 1)
-mapper FILE : path of mapper executable file (default, minimap, only effective with -p option)
-minimap2_sensitive : sensitive mode for minimap2 (default, off; only effective with -p option)
-tmp DIR : directory for temporary files (default .)

Outputs

PREFIX_consensusScaffold.fa
PREFIX_consensusScaffoldComponent.bed

PREFIX is specified by -o

 

Notes

  • Options related to run time Although -t (number of threads) of all commands and -m (memory amount) of the “assemble” command are not mandatory to run, it is recommended to set the values adjusting your machine-environment. These options may severely effect the run time.

    e.g., Available number of threads and memory amount are 12 and 256GB, respectively. -> -t 12 -m 256

  • Minimap2 This tool is used to align PacBio/Oxford-Nanopore long reads and included in Platanus-allee package as the directory of “minimap2-2.0-r191”. When long reads are input through the -p option of “phase” and “consensus” commands, please check Minimap2 is installed as “minimap2” command or specify the path of Minimap2 using the -mapper option.

  • 10X linked-reads input Platanus-allee accepts barcoded fastq files, which have the barcode information in name lines (@…) as the “BX:Z:” tags. These files, “barcoded.fastq”, can be generated using the “longranger basic” command of Long Ranger (https://support.10xgenomics.com/genomeexome/software/pipelines/latest/what-is-long-ranger).

    e.g.

    longranger basic –fastqs=/raw/fastq/directory –id=your_id –sample=your_sample_name
    # Result: your_id/outs/barcoded.fastq.gz

  • Paired-end (mate-pair) input
    The “phase” and “consensus” accept paired-end and/or mate-pair libraries. Paired libraries are classified into “inward-pair” and “outward-pair” according to the sequence direction. For file formats, separate and interleaved files can be input through -IP (-OP) and -ip (-op) options, respectively.

    Inward-pair (usually called “paired-end”, accepted in options “-IP” or “-ip”):

    FWD --->
          5' -------------------- 3'
          3' -------------------- 5'
                                      <--- REV

    Outward-pair (usually called “mate-pair”, accepted in options “-OP” or “-op”):

                                ---> REV
           5' -------------------- 3'
           3' -------------------- 5'
    FWD <---

    Example inputs:

    Inward-pair (separate, insert=300)   : PE300_1.fq PE300_2.fq
    Inward-pair (interleaved, insert=500) : PE500_pair.fq
    Outward-pair (separate, insert=2k)   : MP2k_1.fa MP2k_2.fq

    Corresponding options:

    -IP1 PE300_1_pair.fq PE300_2.fq \
    -ip2 PE500_pair.fq \
    -OP3 MP2k_1.fq MP2k_2.fq

  • Compressed input files Compressed files can be input using the process substitution function (”
    <(command)”) of bash/zsh without temporary uncompressed files.

    e.g.

    # Files compressed by gzip
    platanus_allee assemble -f <(zcat PE_1.fq.gz) <(zcat PE_2.fq.gz)
    # Files compressed by bzip2
    platanus_allee assemble -f <(bzcat PE_1.fq.bz2) <(bzcat PE_2.fq.bz2)