anvi-profile [program]

Creates a single anvi'o profile database. When it is run on a BAM file, depending on the user parameters, the program quantifies coverage per nucleotide position (and averages them per contig), calculates single-nucleotide, single-codon, and single-amino acid variants, as well as structural variants such as insertion and deletions and stores these data into appropriate tables.

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Can provide

single-profile-db misc-data-items-order variability-profile

Can consume

bam-file contigs-db

Usage

This program creates a single-profile-db from a bam-file and contigs-db.

Once you have a single-profile-db, you can run programs like anvi-cluster-contigs, anvi-estimate-metabolism, and anvi-gen-gene-level-stats-databases, as well as use the interactive interface with anvi-interactive. If you want to run these same contigs against multiple BAM files (because you have multiple samples), you’ll combine your single-profile-dbs into a profile-db after you’ve created them all using anvi-merge. See the pages for single-profile-db or profile-db for more you can do with these artifacts.

In short, this program runs various analyses on the contigs in your contigs-db and how they relate to the sample information stored in the bam-file you provided. It then stores this information into a single-profile-db. Specifically, this program calculates

• coverage per nucleotide position (if you’re unsure what coverage refers to, check out this page)
• single-nucleotide, single-codon, and single-amino acid variants (You can find all of those terms on the vocab page linked above, as well as a more detailed explaination here)
• structural variants such as insertions or deletions

Basic Usage

Inputs

This program takes in an indexed bam-file and a contigs-db. The BAM file contains the short reads from a single sample that will be used to create the profile database. Thus, here is a standard run with default parameters:

Alternatively, if you lack mapping data, you can add the flag --blank-profile so that you can still get the functionality of a profile database.

Checking your BAM file: always a good idea

If you want to first check your BAM file to see what contigs it contains, just use the flag --list-contigs to see a comprehensive list.

Profiling a subset of contigs

Note: This describes how to profile a named subset of contigs. To profile a subset of contigs based on their characterists (for example, only contigs of a certain length or that have a certain coverage), see the section below on “contig specifications”

By default, anvi’o will use every contig in your contigs-db. However, if you wish to focus specifically on a subset of these contigs, just provide a file that contains only the names of the contigs you want to analyze, one per line, using the tag --contigs-of-interest.

For example, you could run

Where contigs_i_like.txt looks like this:

SF15-RossSeacontig4922
SF15-RossSeacontig702

Analysis Parameters

Changing these will affect the way that your sequences are analyzed.

Keep in mind that if you plan to merge your resulting single-profile-db with others later in the project, you’ll want to keep these parameters consistent.

Contig Specification

To profile only contigs within a specific length, you can use the flags --min-contig-length and -max-contig-length. By default, the minimum length for analysis is 1000 and there is no maximum length. You can also profile only contigs that have a certain average coverage with the flag --min-mean-coverage.

Specifications for your BAM file

You can also ignore reads in your BAM file with a percent identity to the reference less than some threshold using the flag --min-percent-identity. By default, all reads are used.

For example, the following code will only look at contigs longer than 2000 nts and will ignore BAM file reads with less than 95 percent identity to the reference:

Hierarchical Clustering

To cluster or not to cluster?

By default, anvi’o will not try to cluster your splits (since it takes quite a bit of runtime) unless you are using the tag --blank-profile. If you don’t want to run this, use the tag --skip-hierarchical-clustering.

If you’re planning to later merge this sample with others, it is better to perform clustering while running anvi-merge than at this stage.

However, if you want to bin this single sample or otherwise want clustering to happen, just use the tag --cluster-contigs.

If you do plan to cluster, you can set a custom distance metric or a custom linkage method.

Variability

Anvi-profile will throw away variability data below certain thresholds to reduce noise. After all, if you have a single C read at a position with a 1000X coverage where all other reads are T, this is probably not a variant position that you want to investigate further. By default, it will not analyze positions with coverage less than 10X, and it will further discard variants based on this criteria.

However, you can change the coverage threshold using the --min-coverage-for-variability flag. You can also report every variability position using the flag --report-variability-full.

For example, if you wanted to view every variant, you would profile with the following:

Other Parameters

You should provide the sample name with the flag -S and can provide a description of your project using the --description tag followed by a text file. These will help anvi’o name output files and will show up in the anvi’o interfaces down the line.

You can characterize the codon frequencies of genes in your sample at the cost of some runtime. Despite time being money, codon frequency analysis can be helpful downstream. Simply add the tag --profile-SCVs and watch the magic happen.

If you have prior experience with --profile-SCVs being slow, you will be surprised how fast it is since v6.2

Alternatively, you can choose not to store insertion and deletion data or single nucleotide variant data.

If you know the limits of your system, you can also multithread this program. See the program help menu for more information.

Edit this file to update this information.

Additional Resources

• Another description as part of the metagenomic workflow

Are you aware of resources that may help users better understand the utility of this program? Please feel free to edit this file on GitHub. If you are not sure how to do that, find the __resources__ tag in this file to see an example.