Overview

Teaching: 0 min
Exercises: 20 min

Questions

• How can you obtain and interpret genome assembly information from NCBI?

Objectives

• Practice the usage of command line to get files from the internet and examine their content

• Interpret genome assembly overview from the internet

Genome assembly information

In our exercises, we will examine the quality of assembled Z. tritici genomes that are publicly available. We will use genome assemblies from NCBI. Unfortunately, generating eukaryote de novo genome assembly is beyond the scope of the exercises; in contrast to bacterial genomes, assembly of eukaryotic genomes requires significantly more computational power and time.

The genome assemblies of some Z. tritici isolates downloaded from NCBI GenBank can be found data/fungalgenomics_seidl/assemblies/ (state: 2018). There are continuously more and more genome assemblies being generated and deposited at NCBI.

To get an overview of the number of genome assemblies currently at NCBI. We can obtain an overview file from NCBI with information of all genomes belonging to fungi.

$ wget https://ftp.ncbi.nlm.nih.gov/genomes/genbank/fungi/assembly_summary.txt

Exercise

How many fungal genomes are deposited in the database today?

Solution

cat assembly_summary.txt | grep -v "^#" | wc -l

Inspect the assembly_summary file further. You can look at README_assembly_summary.txt for a description of the columns.

Exercise

What does the term ‘representative genome’ refer to?

Solution

Representative genomes: Additional high-quality genomes are identified by clustering genomes and applying weighting metrics that include consideration of species-level taxonomic classification (e.g., a preference for type strain) and assembly quality (e.g. a preference for complete genomes but WGS is allowed). Additional quality assurance analysis is being added to add consideration of annotation quality metrics such as assessing the number of frame-shifted proteins (compared to close neighbours), presence of the set of expected rRNA and tRNAs, and gene density. We also take into consideration taxonomic diversity and will include some genomes that are taxonomic outliers for which little functional information is available in the representative genome collection.

We can now try to find out for how many different species there is at least one genome sequenced. We can do this using the command line.

$ cat assembly_summary.txt | grep -v "^#" | cut -f8 | cut -f1,2 -d" " | sort -u | wc -l

With grep you can obtain the lines matching a specific expression. The -v inverts the matches, i.e., it will return all lines that do not match the expression. In our case, we are excluding linkes starting with a # as these are comment lines. We then obtain the eights column - why? (Tip: Take a look at the information stored in each column) with cut -f 8 and the only retain the first two entries separated by a space -d " ". Lastly, we make these results unique and then count the number of lines.

Exercise

Can you modify the code to obtain the code to obtain the fungal genus for which most genome assemblies have been produced so far? Explain the logic of the code.

Solution

cat assembly_summary.txt | grep -v "^#" | cut -f8 | cut -f1,1 -d " " | sort | uniq -c | sort -k1,1nr | head. The logic is similar to above but we only keep the genus information (first entry in the second cut), and then sort the output numerically to get the highest number on top of the list.

We can now use a very similar code to obtain the number of Zymoseptoria genome assemblies currently available.

$ cat assembly_summary.txt | grep -v "^#" | cut -f8 | cut -f1,2 -d" " | sort | uniq -c | sort -k1,1nr | grep "Zymoseptoria"

Exercise

Can you think about a code that would give you the number of complete (chromosome-level) genome assemblies?

Key Points

• You can obtain assembly information from NCBI

• You can subset this information and determine the number of genome assemblies