Pipeline which make phylogeny with sequence of different sample
Technician, François HIRIART
Hospital Engineers, Aurelien BIRER
Professor, Richard BONNET
Phylosnip is a pipeline of bacterial typing. The pipeline use the data of high-throughput sequencing which will be mapped to an haploid reference genome. Next, Phylosnip find SNP, indels and MNP to discriminate sample to a core genome but also between themselves. Finally, Phylosnip will produce a distance matrice and a network graph.
This will install the repositories on github.
cd where/you/want/to/install
git clone https://github.com/Frahiriart/Phylosnip.git
The script "setup.sh" will install all binaries of this program.
cd where/you/want/to/install/Phylosnip
chmod u+x setup.sh
./setup.sh
- a reference genome in FASTA or GENBANK format (can be in multiple contigs)
- sequence read files in FASTQ or FASTA format (can be .gz compressed) format
- 1 folder per sample, named by the name of the sample
- 1 folder which collect and compare the SNP result of all strain and this folder have also distance matrix and network phylogeny, named merge_genome_core_result.
This table come from snippy page
| Extension | Description |
|---|---|
| .tab | A simple tab-separated summary of all the variants |
| .csv | A comma-separated version of the .tab file |
| .html | A HTML version of the .tab file |
| .vcf | The final annotated variants in VCF format |
| .bed | The variants in BED format |
| .gff | The variants in GFF3 format |
| .bam | The alignments in BAM format. Includes unmapped, multimapping reads. Excludes duplicates. |
| .bam.bai | Index for the .bam file |
| .log | A log file with the commands run and their outputs |
| .aligned.fa | A version of the reference but with - at position with depth=0 and N for 0 < depth < --mincov (does not have variants) |
| .consensus.fa | A version of the reference genome with all variants instantiated |
| .consensus.subs.fa | A version of the reference genome with only substitution variants instantiated |
| .raw.vcf | The unfiltered variant calls from Freebayes |
| .filt.vcf | The filtered variant calls from Freebayes |
| Name | Description |
|---|---|
| CHROM | The sequence the variant was found in eg. the name after the > in the FASTA reference |
| POS | Position in the sequence, counting from 1 |
| TYPE | The variant type: snp ins del complex |
| REF | The nucleotide(s) in the reference |
| ALT | The alternate nucleotide(s) supported by the reads |
| QUAL | probability that the ALT allele is incorrectly specified, expressed on the the phred scale (-10log10(probability)). |
| FILTER | Either "PASS" or a semicolon-separated list of failed quality control filters. |
| INFO | additional information (TYPE=Variant_Type;DP=Depth;VD=number_of_Variant;AF=Frequence_of_Variant). |
| Type | Name | Example |
|---|---|---|
| SNV | Single Nucleotide Variant (=SNP) | A => T |
| MNV | Multiple Nuclotide Polymorphism | GC => AT |
| Insertion | Insertion of Nucleotide | ATT => AGTT |
| Deletion | Deletion of Nucleotide | ACGG => ACG |
| Complex | Combination of snp/mnp | ATTC => GTTA |
- a set of Snippy folders which used the same reference sequence (
--genome).
| Extension | Description |
|---|---|
| .aln | A core SNP alignment in the FASTA format |
| .full.aln | A whole genome SNP alignment (includes invariant sites) |
| .tab | Tab-separated columnar list of core Variant sites with alleles and annotations |
| .nway.tab | Tab-separated columnar list of all Variant sites with alleles and annotations |
| .vcf | Multi-sample VCF file with genotype GT tags for all discovered alleles |
| .txt | Tab-separated columnar list of alignment/core-size statistics |
| _density_filtered_keep.vcf | Tab-separated columnar list of core Variant sites with alleles and annotations which are filtered by density |
| _density_filtered_unkeep.vcf | Tab-separated columnar list of core Variant sites with alleles and annotations which are reject after the density filter |
| _density_filtered_keep_SNP_dist.tsv | Distance Matrice of all sample between themselves |
| SNP_network | Phylogeny Network |
if you want to test Pylosnipping with data test you must have SRA toolkit. You can download SRA toolkit with this command.
wget http://ftp-trace.ncbi.nlm.nih.gov/sra/sdk/2.4.1/sratoolkit.2.4.1-ubuntu64.tar.gz
tar xzvf sratoolkit.2.4.1-ubuntu64.tar.gz
Data which will be download come from this
cd where/you/want/to/install/Phylosnip/test
for i in `cat SRR_Acc_List.txt`; do ~/where/is/sratoolkit.2.9/bin/fastq-dump --split-files $i; gzip -9 $i*; done
sudo apt install rename
for b in `awk '{print "s/"$11"/"$8"/";}' SraRunTable.txt`;do rename `echo $b` *; done
wget https://www.ncbi.nlm.nih.gov/sviewer/viewer.cgi?tool=portal&save=file&log$=seqview&db=nuccore&report=fasta&id=378697983&
cd where/you/want/to/install/Phylosnip/test
/fastq2phylotreeV1.py -input test -g test/sequence.fasta -o where/you/want/your/resut
- Java = 1.8
- Perl >= 5.12
- R >= 3.2.5
- Python 3.6
- Perl Modules : bioperl >= 1.6
- snippy >= 4.3.5
- picard.jar >= 2.18.8
- GenomeAnalysisTK.jar >= 4.0.11.0
- samtools >= 1.7
- bwa mem >= 0.7.12
- bcftools >= 1.7
- GNU parallel >= 2013xxxx
- snpEff >= 4.3
- bedtools >= 2.0
- bcftools >= 1.7
- minimap2 >= 2.0
- vcflib >= 1.0 (vcfstreamsort, vcfuniq, vcffirstheader)
- snp-sites >= 2.0
- seqtk >= 1.2
- samclip >= 0.2
- readseq >= 2.0
- vt >= 0.5
- vcflib >= 1.0
For Linux (compiled on Ubuntu 16.04 LTS) some of the binaries, JARs and scripts are included.
And the binaries can be install with the file setup.sh.