seung1yoo / phyne

phyne (PHYlogenetic analysis pipeliNE)

Workflow

phyne는 phylogenetic analysis를 위한 파이프라인으로 다양한 인풋을 지원합니다.

Development Environment

• server : wolf
• language : python 2.7

Dependents

Common dependents

• MAFFT v7.123b
• MUSCLE v3.8.31
• Gblocks 0.91b
• FastTree 2.1.11
• R package (RColorBrewer, ggplot2, phylogram)

MLST dependents

• mlst
• PubMLST

ortholog dependents

• OrthoMCL v2.0.9.M
• BLAST 2.2.26

Usage

Help message

$python bin/phyne.py --help
usage: phyne.py [-h] [--mode {mlst,nssnp,ortholog}] [--config CONFIG]
                [--outdir OUTDIR] [--prefix PREFIX]

optional arguments:
  -h, --help            show this help message and exit
  --mode {mlst,nssnp,ortholog}
  --config CONFIG
  --outdir OUTDIR
  --prefix PREFIX

MLST

Workflow

General Command line

$python bin/phyne.py --mode mlst --config [mlst.conf] --outdir [result] --prefix [testset]

Input & Output

• Input : Bacterial genome sequences (FASTA format)

• Output

1. {outdir}/{prefix}.mlst_profile.xls : MLST profile (a tab-separated line)

GCA_000439795.1 aphagocytophilum        64      pheS(42)        glyA(32)        fumC(28)        mdh(18) sucA(40)        dnaN(28)      atpA(26)
GCA_000013125.1 aphagocytophilum        161     pheS(42)        glyA(32)        fumC(28)        mdh(18) sucA(82)        dnaN(28)      atpA(26)
GCA_000439775.1 aphagocytophilum        64      pheS(42)        glyA(32)        fumC(28)        mdh(18) sucA(40)        dnaN(28)      atpA(26)
GCA_000689655.1 aphagocytophilum        215     pheS(103)       glyA(77)        fumC(28)        mdh(4)  sucA(94)        dnaN(28)      atpA(60)
GCA_000689635.2 aphagocytophilum        82      pheS(3) glyA(33)        fumC(29)        mdh(3)  sucA(2) dnaN(2) atpA(4)
GCA_000964685.1 aphagocytophilum        64      pheS(42)        glyA(32)        fumC(28)        mdh(18) sucA(40)        dnaN(28)      atpA(26)
GCA_000689615.1 aphagocytophilum        217     pheS(104)       glyA(78)        fumC(70)        mdh(53) sucA(95)        dnaN(77)      atpA(1)
GCA_000478425.1 aphagocytophilum        64      pheS(42)        glyA(32)        fumC(28)        mdh(18) sucA(40)        dnaN(28)      atpA(26)
GCA_000968455.1 aphagocytophilum        -       pheS(42)        glyA(32)        fumC(28)        mdh(18?)        sucA(40)        dnaN(28)       atpA(26)

• col 1 : the genome name
• col 2 : the matching PubMLST scheme name
• col 3 : the ST (sequence type)
• col 4 ~ : the allele IDs

2. {outdir}/{prefix}.mlst_sequence.fa : Allele sequences are arranged in order
3. {outdir}/{prefix}.mlst_sequence.fa.order : THE order
4. {outdir}/{prefix}.mlst_align.fa : multiple sequence alignment (fasta)
5. {outdir}/{prefix}.mlst_align.phylip : multiple sequence alignment (phylip)
6. {outdir}/{prefix}.mlst_align.newick : tree (newick)
7. {outdir}/{prefix}.mlst_align.dist : Sequence distance metrics
8. {outdir}/{prefix}.PhylogeneticTree.png : very simple phylogenetic tree figure
9. {outdir}/{prefix}.PCA.png : PCA plot

Config file

• Without target scheme

$ cat bin/phyne_mlst.conf
{
        "target_scheme" : "",
        "input_genome" : {
                "mygenome" : "denovogenome.fa",
                "relativegenome1" : "Salmonella_enterica_subsp_enterica_serovar_Typhimurium_DT104_v1.fa",
                "relativegenome2" : "Salmonella_enterica_subsp_enterica_serovar_Typhi_str_CT18_v1.fa",
                "relativegenome3" : "Salmonella_enterica_subsp_enterica_serovar_Weltevreden_str_10259_v0.2.fa"
        },
        "phyne_mlst_exe" : "/BiO/BioPeople/siyoo/phyne/bin/phyne_mlst.py"
}

• With target scheme Scheme list

$ cat bin/phyne_mlst.conf
{
        "target_scheme" : "aphagocytophilum",
        "input_genome" : {
                "genome1" : "GCA_000013125.1_ASM1312v1_genomic.fna",
                "genome2" : "GCA_000439775.1_ASM43977v1_genomic.fna",
                "genome3" : "GCA_000439795.1_ASM43979v1_genomic.fna",
                "genome4" : "GCA_000478425.1_ASM47842v1_genomic.fna",
                "genome5" : "GCA_000689655.1_MRK1.0_genomic.fna"
        },
        "phyne_mlst_exe" : "/BiO/BioPeople/siyoo/phyne/bin/phyne_mlst.py"
}

NSSNP (Non-Synoymous SNP)

Workflow

multisample vcf ==> Defind and select Non-Synoymous SNPs ==> Phylogenetic analysis

General Command line

$python bin/phyne.py --mode nssnp --config [mlst.conf] --outdir [result] --prefix [testset]

Input & Output

• Input file : Multisample variant calling data (format : vcf)
• Output file

1. {outdir}/{prefix}.nssnp.fasta : Nonsynoymous SNP concatenated data (format : fasta)
2. {outdir}/{prefix}.nssnp.newick : tree (format : newick)
3. {outdir}/distance.txt : Sequence distance metrics
4. {outdir}/PhylogeneticTree.png : Phylogenetic tree figure
5. {outdir]/PCA.png : PCA plot

Config file

### REQUERMENT ARGUMENTS ###
INPUT=path/to/multisample.snpeff.test.vcf
SAMPLE_IDS=SAMPLE1,SAMPLE2,SAMPLE3,SAMPLE4,SAMPLE5

### OPTIONAL ARGUMENTS ###
GQ=60
DP=10
REF=90.0
HET=70.0,30.0
ALT=90.0

Mandatory arguments

• config (file_path) : Pipeline usage input configure file
• outdir (dir_path) : Output directory
• prefix (STRING) : Output result file name

Filter optional arguments

• GQ (INT) : Genotype quality score (default=60)
• DP (INT) : Genotype total read depth (default=10)
• REF (FLOAT) : Reference allele ratio (default=90.0)
• HET (FLOAT,FLOAT) : Heterozygous allele ratio (default=70.0,30.0 (mean = 70 : 30))
• ALT (FLOAT) : Alternative allele ratio (default=90.0)

! The above options will be removed if they do not SNP the conditions.

Ortholog (Single copy genes)

Based on the evolutionary similarity of sequences, a pipeline that draws phylogenetic trees by selecting only single copy genes of two or more species/genomes

Workflow

• Step1. Clustering ortholog Groups of Protein Sequences with orthoMCL
• Step2. Single copy gene selection
• Step3. Multiple sequence alignment
• Step4. Remove ambiguously aligned regions
• Step5. Merge clean alignment and Make tree
• Step6. Calculate the sequence distance

General Command line

python bin/phyne.py --mode ortholog --config [ortholog.conf] --outdir [result] --prefix [testset]

Input & Output

• Input : protein sequences (FASTA format)

• Output : Intermediate directory

1. {outdir}/1.orthoMCL : orthoMCL-Run Result files
2. {outdir}/2.mafft : multiple sequence alignment of each single copy gene cluster (fasta)
3. {outdir}/3.Gblock : The sequence from which the region with low quality is removed (fasta)
4. {outdir}/4.fasttree : newick file

• Ooutput : Final files

1. {outdir}/Report/ortholog.fa : multiple sequnece alignmet merge
2. {outdir}/Report/newick.txt : tree
3. {outdir}/Report/SinglecopyGene.xls : single copy gene lists between species
4. {outdir}/Report/distance.txt : Sequence distance metrics
5. {outdir}/Report/PhylogeneticTree.png : very simple phylogenetic tree figure
6. {outdir}/Report/PCA.png : PCA plot

Config file

• phyne_ortholog.conf :

#otrhoMCL
##########################################################################
orthoMCL        =       Y        
ortho_fasta_dir =       /BiO/BioPeople/boram/Projects/Phylogenetic/pepiteds
ortho_thread    =       40
parser  =       Y
ortholog_fa     =       Y
##########################################################################
Tree    =       fasttree

• orthoMCL을 실행 시키려면 Y, 다음 Step만 진행 하고자 한다면 N
• ortho_fasta_dir은 종들의 amino acid sequence (fasta) 파일이 있는 directory 경로 입력
• ortho_thread는 orthoMCL의 thread를 설정해주는 것으로, 기본 40으로 설정
• parser는 Single copy gene을 선별하는 script 실행, 이미 진행 했다면 N으로 설정
• ortholog_fa는 종 별 Single copy gene의 서열을 모아놓은 fasta 생성

! 처음 실행시킬때에는 모두 Y로 실행 시키고, 원하는 step부터 실행시키고자 할 때 N으로 변경하여 사용하면 된다.

Contributors

• [email protected]
• [email protected]
• [email protected]

lastest update : 2019-05-24