refineM

安装需要的相关软件

##numpy
sudo apt install python-numpy
##Prodigal
git clone [https://github.com/hyattpd/Prodigal.git](https://github.com/hyattpd/Prodigal.git)
cd Prodigal/
make install
##blast+
apt install ncbi-blast+
##diamond
wget [http://github.com/bbuchfink/diamond/releases/download/v0.9.24/diamond-linux64.tar.gz](http://github.com/bbuchfink/diamond/releases/download/v0.9.24/diamond-linux64.tar.gz)
tar xzf diamond-linux64.tar.gz
cp diamond /usr/local/bin/
##krona
git clone [https://github.com/marbl/Krona.git](https://github.com/marbl/Krona.git)
cd Krona/KronaTools/
cd /home/llt/database/krona
mkdir taxonomy
sudo ./install.pl --taxonomy /home/llt/database/krona/taxonomy
sudo apt install curl
./updateTaxonomy.sh
./updateAccessions.sh

安装refineM

sudo apt install python-pip
sudo pip install refinem
sudo pip install pysam dendropy -i https://pypi.mirrors.ustc.edu.cn/simple/
sudo apt install python-scipy
sudo pip install matplotlib==1.5.3 ##refineM与2.0以上版本的matplotlib不兼容

使用

Removing contamination based on genomic properties

##refinem scaffold_stats -c 16    
refinem scaffold_stats -c 8 -x fasta /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/contigs.fasta /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/maxbin/1000 stats_out /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/ssg1_30m.sorted.bam

##refinem outliers /scaffold_stats.tsv 
refinem outliers stats_out/scaffold_stats.tsv outlier_out

##refinem filter_bins  /outliers.tsv 
refinem filter_bins -x fasta /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/maxbin/1000 outlier_out/outliers.tsv filtered_out

Removing contamination based on taxonomic assignments

##refinem call_genes -c 40  
refinem call_genes -c 8 -x fasta /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/maxbin/1000/filtered_out gene_out

下载refineM所需的数据库
https://data.ace.uq.edu.au/public/misc_downloads/refinem/gtdb_r86_protein_db.2018-09-25.tar.gz
https://data.ace.uq.edu.au/public/misc_downloads/refinem/gtdb_r86_taxonomy.2018-09-25.tsv

##建立dmnd数据库供refineM使用
diamond makedb --in gtdb_r86_protein_db.2018-09-25.faa -d gtdb_r86_protein_db.2018-09-25 
##refinem taxon_profile -c 40  /scaffold_stats.tsv   
refinem taxon_profile -c 8 gene_out stats_out/scaffold_stats.tsv /home/llt/database/refinem/gtdb_r86_protein_db.2018-09-25.dmnd /home/llt/database/refinem/gtdb_r86_taxonomy.2018-09-25.tsv taxon_profile_out

##refinem taxon_filter -c 40  taxon_filter.tsv
refinem taxon_filter -c 8 /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/maxbin/1000/taxon_profile_out taxon_filter.tsv

##refinem filter_bins  taxon_filter.tsv 
refinem filter_bins -x fasta  filtered_out/  taxon_filter.tsv taxon_filtered_out

Removing contigs with incongruent 16S rRNA genes (requires version >=0.0.22)

diamond makedb --in bac_arc_ssu_r86.fna -d bac_arc_ssu_r86
sudo apt install hmmer
##refinem ssu_erroneous     
refinem ssu_erroneous -x fasta /home/llt/experiment/data/clean/subsamble_30m/assembly_spades/maxbin/1000/  taxon_profile_out/ /home/llt/database/refinem/bac_arc_ssu_r86.dmnd /home/llt/database/refinem/bac_arc_taxonomy_r86.tsv ssu_output_dir