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**BUSCO: Benchmarking Universal Single-Copy Orthologs**

Documentation by Dayana Salas-Leiva (last update 04-13-2020)

Web:http://busco.ezlab.org/

You can check the completeness of your genome by picking out single-copy orthologs. **BUSCO** runs **tBLASTn**, **AUGUSTUS**, and **HMMER 3** based on single-copy orthologs from the **OrthoDB** database.

There are two main dataset versions of busco: **odb9** (contains 303 orthologs only compatible with Busco3) and **odb10** (contains 255 orthologous only compatible with Busco4)

The following are examples of a shell script for genomic and a proteomic search:

***  BUSCO 3 ***

** Genomic: **

#!/bin/bash
   #$ -S /bin/bash
   ./etc/profile
   #$ -pe threaded 1
   #$ -cwd
   cd $PWD
   #busco3.0.0
   source activate busco-3
   export AUGUSTUS_CONFIG_PATH="/home/dsalas/Shared/BUSCO/config"
   run_BUSCO.py -i <fasta_file> -o <output_dir_name> -l /home/dsalas/Shared/BUSCO/eukaryota_odb9 -m geno --cpu 1
   conda deactivate

** Proteomic: **

#!/bin/bash
   #$ -S /bin/bash
   ./etc/profile
   #$ -pe threaded 1
   #$ -cwd
   cd $PWD
   #busco3.0.0
   source activate busco-3
   export AUGUSTUS_CONFIG_PATH="/home/dsalas/Shared/BUSCO/config"
   run_BUSCO.py -i <fasta_file> -o <output_dir_name> -l /home/dsalas/Shared/BUSCO/eukaryota_odb9 -m prot --cpu 1
   conda deactivate

** * BUSCO 4.0.5 * **

** Genomic: **
   #!/bin/bash
   #$ -S /bin/bash
   ./etc/profile
   #$ -pe threaded 1
   #$ -cwd
   cd $PWD
   source activate busco
   #BUSCO 4.0.5
   export AUGUSTUS_CONFIG_PATH="/home/dsalas/Shared/BUSCO/config"
   busco -i <input_scaffolds_file> -o <output_dir_name> -l /home/dsalas/Shared/BUSCO/eukaryota_odb9 -m geno --cpu 1

**Proteomic:**

#!/bin/bash
   #$ -S /bin/bash
   . /etc/profile
   #$ -pe threaded 1
   #$ -cwd
   cd $PWD
   source activate busco
   #BUSCO 4.0.5
   export AUGUSTUS_CONFIG_PATH="/home/dsalas/Shared/BUSCO/config"
   busco -i <predicted protein fasta> -o <output_dir-name> -l /home/dsalas/Shared/BUSCO/eukaryota_odb9 -m prot --cpu 1

**Note**: Take out the mitochondrial genome before running this analysis.

**In the output folder:**

-3 folders for the outputs of tBLASTn, AUGUSTUS, and HMMER, and a folder containing the single-copy BUSCO sequences. If you want to look into the details of the amino acid or nucleotide sequences that were predicted by AUGUSTUS and align them manually (for example, against a transcript contig), pick out the .faa or .fna files from the directory augustus_output/extracted_proteins

-"short summary" file showing how many BUSCO genes were found, how many were complete, fragmented, etc.

-"full table.tsv" showing a list of your contigs matched to BUSCO IDs.

-"missing busco list" showing a list of BUSCO genes that were not found in your genome.

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