Assembly

Assembly

Use the most popular assembler SPAdes to assemble a bacterial genome.

Installation

conda install spades

Assembly

When using SPAdes it is typical to choose at least 3 k-mer sizes. One low, one medium and one high. We will use 33, 55 and 91.

First merge the unpaired reads

cd seqtk
cat SRR10561173_2U_seqtk.fastq.gz >> SRR10561173_1U_seqtk.fastq.gz
rm SRR10561173_2U_seqtk.fastq.gz
mv SRR10561173_1U_seqtk.fastq.gz SRR10561173_U_seqtk.fastq.gz
cd ..

then run spades

spades.py -1 ~/seqtk/SRR10561173_1P_seqtk.fastq.gz -2 ~/seqtk/SRR10561173_2P_seqtk.fastq.gz -s ~/seqtk/SRR10561173_U_seqtk.fastq.gz -k 33,55,91 -o ~/assembly

Check the result.

File	content
contigs.fasta	assembled contigs
scaffolds.fasta	assembled scaffolds
assembly_graph.fastg	the graph of contigs
assembly_graph_with_scaffolds.gfa	the graph of scaffolds
spades.log	the log file of the spades running

Quanlity assessment of the assembly

• Install quast and bandage

  quast have some compatibility problem with the bioinfo environment, the solution is to install it in a separate environment.

    conda create -n quast
    conda activate quast
    conda install quast
  

• Run quast

  quast use a reference to assess the quality of the assembly. Upload the reference genome and annotation file salmonella_typhimurium_lt2.fasta and salmonella_typhimurium_lt2.gff to the server.

    quast ~/assembly/SRR10561173.fasta -R ~/seq/salmonella_typhimurium_lt2.fasta -G ~/seq/salmonella_typhimurium_lt2.gff -o ~/quast/
  

  Check the result.

• Run bandage

  Download and install bandage from bandage website, or copy from my folder.

  Download the FASTG graph file of SPAdes to your laptop, open it with bandage.