• python 3 or greater
• Biopython
• trimmomatic
• prinseq
• deconseq
• bbmerge
• blastn
• fastq-pair

java -jar ~/Trimmomatic-0.36/trimmomatic-0.36.jar PE $FILEF $FILER paired_output_${FILEF%} \
unpaired_output_${FILEF%} paired_output_${FILER%} unpaired_output_${FILER%} -trimlog output.log \
ILLUMINACLIP:TruSeq3-PE-2.fa:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:60

perl ~/prinseq-lite-0.20.4/prinseq-lite.pl -verbose -derep 1245 -lc_method entropy \
-lc_threshold 50 -trim_qual_right 20 -trim_qual_left 20 -trim_qual_type mean \
-trim_qual_rule lt -trim_qual_window 2 -trim_qual_step 1 -trim_tail_left 5 \
-trim_tail_right 5 -min_len 60 -min_qual_mean 25 -ns_max_p 1 \
-fastq $FILEA -fastq2 $FILEB -log ${FILEA%}_prinseq.log -out_bad ${FILEA%}_prinseq_bad \
-out_good ${FILEA%}_prinseq_good

perl deconseq-standalone-0.4.3/bwasw_modified_source/deconseq.pl \
   -f file.fq \
   -dbs hs_ref_GRCh37_p7 \
   -dbs_retain virus -out_dir deconseq/ \
   -id $FILE -i 90 -c 95 -group 1

fastq_pair $FILEF.fastq $FILER.fastq

~/bbmap/bbmerge.sh 
in1=$FILEF \ 
in2=$FILER out=${FILEO%}_merged.fa \
outu=${FILEO%}_unmereged.fa hist=${FILEO%}_histogram.txt

xtract_forward allows you to extract your unmerged high quality R1 reads.

After merging your paired end reads programs typically give you 1 R1 and R2 merged file and 1 file that contained all your R1 an R2 reads that did not merge. If doing a simple classification (e.g., Blast) you could end up throwing away useful R1 data from your analysis. xtract-forward allows you to extract all your R1 reads from your unmerged file so you can then concatenate it with your R1/R2 merged file.

Will need a file that only contains the accession numbers in your fna file. Below is an example of how you can get that from your fna file:

sed 's/\s.*$//' subset_viral_genomic.fna | grep ">" | sed 's/>//' | awk '{print $1}' > subset_viral_accesion.txt

Script allows you to change the headers of fna files downloaded and concatenated from NCBI ref seq. After this script headers should include nucleotide Accession number followed by full taxonomic lineage

python get_taxlineage_fna.py --acc test/subset_viral_accesion.txt --input test/subset_viral_genomic.fna --output test/viral_subset.genomic.new.header.fna

ncbi-blast-2.6.0+/bin/blastn \
  -query File1.fasta \
  -outfmt "6 stitle" -out File1.blastn.txt \
  -db viral -dust no \
  -max_target_seqs 1 -perc_identity 90 -evalue 0.0001 -min_raw_gapped_score 105