Single_Genome_Assembly_script_23.08.2021.sh

## Quality Control

trimmomatic PE -threads 4 -phred33 *R1* *R2* PF1.fq UF1.fq PF2.fq UF2.fq ILLUMINACLIP:NexteraPE-PE.fa:2:30:10 LEADING:20 TRAILING:20 MINLEN:50

cat PF1.fq UF1.fq > forward.fq
cat PF2.fq UF2.fq > reverse.fq

rm PF1.fq
rm UF1.fq
rm PF2.fq
rm UF2.fq


## Dereplication


seqkit rmdup forward.fq -s -o DPF1.fq -j 4
seqkit rmdup reverse.fq -s -o DPF2.fq -j 4

rm forward.fq
rm reverse.fq


## Deconvolution


bbduk.sh in=DPF1.fq out=forward.fq  ref=phiX/phi_X174_phage.fa k=31 hdist=1
bbduk.sh in=DPF2.fq out=reverse.fq  ref=phi_X174_phage.fa k=31 hdist=1

rm DPF1.fq
rm DPF2.fq


## Pairing sequences (paired & unpaired)


fastq_pair forward.fq reverse.fq

rm forward.fq
rm reverse.fq

cat *single.fq > unpaired.fq
rm *single.fq


## Denovo Assembly using (i) Illumina


spades.py --pe1-1 forward.fq.paired.fq --pe1-2 reverse.fq.paired.fq --pe1-s unpaired.fq -o spades_folder_illumina -t 4 -m 7 --only-assembler


## Selecting long scaffolds (i) Illumina


seqkit seq spades_folder_illumina/scaffolds.fasta -m 2000 -g > scaffolds_2k_illumina.fasta


## results (i) Illumina


grep ">" scaffolds_2k_illumina.fasta


## Quality Control for ONT reads


cat BC10.fastq | NanoFilt -q 7 -l 500 --readtype 1D > BC10_HQ.fq


## Denovo Assembly using (ii) Illumina hybrid ONT


spades.py --pe1-1 forward.fq.paired.fq --pe1-2 reverse.fq.paired.fq --pe1-s unpaired.fq -o spades_folder_hybrid   -t 4 -m 7 --only-assembler  --nanopore BC10_HQ.fq


## Selecting long scaffolds (ii) Illumina hybrid ONT


seqkit seq spades_folder_hybrid/scaffolds.fasta   -m 2000 -g > scaffolds_2k_hybrid.fasta


## results (ii) Illumina hybrid ONT


grep ">" scaffolds_2k_hybrid.fasta


## Cleaning


rm *fq


## Comparing Assemblies


makeblastdb -in scaffolds_2k_hybrid.fasta -dbtype nucl
blastn -query scaffolds_2k_illumina.fasta -db scaffolds_2k_hybrid.fasta -outfmt 6 -evalue 10e-5 -num_threads 4 -strand both -subject_besthit  > OutSelfBlast.txt