Custom scripts to interact with the Logan dataset.

Logan is a dataset comprising DNA and RNA sequences derived from a December 2023 snapshot of the entire NCBI Sequence Read Archive through genome assembly. It offers two sets of related sequences: unitigs and contigs. Unitigs retain nearly all original sample information, while contigs minimize variation to increase sequence lengths. Both datasets are available on a public S3 bucket provided by the Registry of Open Data at AWS, in compressed form. Downloading either unitigs or contigs enables users to access the extensive information within the SRA 10x (40x for contigs) more efficiently in terms of time and disk space compared to raw reads, albeit with minor sensitivity loss and enhanced contiguity.

For more informations : https://github.com/IndexThePlanet/Logan

Requirements:

Prior to using this script, ensure you have installed the following:

• aws CLI
• rcgrep (v0.1.0)
• zstd (v1.5.5)
  • For mapping :
• minimap2
• samtools

Note: If you're using the I2BC cluster, you need to connect to the node to access all these tools.

To install aws CLI, please follow the instractions via :

https://docs.aws.amazon.com/cli/latest/userguide/getting-started-install.html

To install rcgrep please execute the command line

pip install rcgrep

To install zstd please execute the command line

We recommand installing zstd via conda.
conda install anaconda::zstd

To install minimap2, please follow the instruction:

https://github.com/lh3/minimap2?tab=readme-ov-file#install

Cloning

Execute the command line :

• git clone https://github.com/Fadwa7/LoganScripts.git && cd LoganScripts

Tutorial :

In this example we download unitigs for 5 pancreatic cancer samples and search them for KRAS oncogenic variants using 33-nt sequence queries. The data used in this tutorial is present in data

Downloading files using id_accessions.txt:

To execute download.sh script, you'll need two files: id_accessions.txt and a directory

• id_accessions.txt: contains SRR Identifiers for your samples, with each ID on a separate line.
• directory : The absolute path to the output directory, e.g : . (current directory)
• data_type : desired data type : contigs or unitigs.

Execute the following command:

./download.sh data/id_accessions.txt . unitigs

Output :

ERR1880167.unitigs.fa.zst
ERR1880168.unitigs.fa.zst
ERR1880169.unitigs.fa.zst
ERR1880170.unitigs.fa.zst
ERR1880172.unitigs.fa.zst

Search for queries:

To execute search.sh script, you'll need two files: paths.txt and query.fa

• paths.txt: contains absolute path to your samples (.zst or .fa)
• query.fa: contains queries in fasta format. (make sure you use .fasta or .fa extension).
  N.B: you can use a simple file instead of the fasta format, with each query on its own line.

Execute the following command:

./search.sh data/paths.txt data/query.fa

In the terminal you should have this output

List of queries:
ACTCTTGCCTACGCCATCAGCTCCAACTACCAC
ACTCTTGCCTACGCCAACAGCTCCAACTACCAC
CTCTTGCCTACGCCACAAGCTCCAACTACCACA
CTCTTGCCTACGCCACGAGCTCCAACTACCACA
GGCACTCTTGCCTACGTCACCAGCTCCAACTAC
ACTCTTGCCTACGCCAGCAGCTCCAACTACCAC
Searching for queries...

~~~~~~ Query result for ERR1880167.unitigs.fa.zst ~~~~~~~

>ERR1880167_199416 ka:f:41.6   L:-:1359598:+ L:-:1660377:+  L:+:4332788:+ 
AGGCCTGCTGAAAATGACTGAATATAAACTTGTGGTAGTTGGAGCTGTTGGCGTAGGCAAGAGTGCCTTGACGATACAGCTAATTCAGAATCATTTTGTGGACGAATATGATCCAACAATAGAGGATTCCTACAGGAAGCAAGTAGTAAT

~~~~~~ Query result for ERR1880168.unitigs.fa.zst ~~~~~~~

>ERR1880168_1809982 ka:f:34.8   L:+:2372536:+  L:-:1037400:+ L:-:1042496:+ 
AGGCCTGCTGAAAATGACTGAATATAAACTTGTGGTAGTTGGAGCTGTTGGCGTAGGCAAGAGTGCCTTGACGATACAGCTAATTCAGAATCATTTTGTGGACGAATATGATCCAACAATAGAGGATTCCTACAGGAAGCAAGTAGTAATTGATGGAGAAACCTGTCTC

~~~~~~ Query result for ERR1880169.unitigs.fa.zst ~~~~~~~

>ERR1880169_1443447 ka:f:31.3   L:-:2508470:-  L:+:1352292:- 
CTGAATATAAACTTGTGGTAGTTGGAGCTGATGGCGTAGGCAAGAGTGCCTTGACGATACA

~~~~~~ Query result for ERR1880170.unitigs.fa.zst ~~~~~~~

>ERR1880170_2665195 ka:f:29.5   L:-:2236668:-  L:+:251215:- 
CTGAATATAAACTTGTGGTAGTTGGAGCTGATGGCGTAGGCAAGAGTGCCTTGACGATACA

~~~~~~ Query result for ERR1880172.unitigs.fa.zst ~~~~~~~

>ERR1880172_9459260 ka:f:168.9   L:-:3582700:+  L:+:6737383:+ 
CTGAATATAAACTTGTGGTAGTTGGAGCTGATGGCGTAGGCAAGAGTGC

Process completed successfully.

Mapping

mapping.sh is a script which aims to download all contigs from a pre-defined list of accessions, then map these contigs on the fly to a gene of interest using minimap2.

To execute this script, you'll need to :

• id_chicken_avail.txt: id accessions that contain chickens genomes
• mc1r.fa : gene from the chicken reference genome
• output_directory: output directory where storing mapping result.

Execute the following command:

./mapping.sh data/id_chicken_avail.txt data/mc1r.fa .

At the end, you should have mapping outputs in your output directory :

SRR20750026.minimap2_output
SRR20750040.minimap2_output
SRR20750042.minimap2_output

If you have any suggestions, please feel free to reach out to us at [email protected].