TIRmite

Build and map profile Hidden Markov Models for Terminal Inverted Repeat families (TIR-pHMMs) to genomic sequences for annotation of MITES and complete DNA-Transposons with variable internal sequence composition.

If you have a draft TE model (i.e. from RepeatModeler or EDTA) and want to identify the TIR's to use with TIRmite - we recommend using tSplit a tool for extraction of terminal repeats from complete transposons.

Table of contents

• About TIRmite
• Algorithm overview
• Options and usage
  • Installing TIRmite
  • Example usage
  • Standard options
  • Custom DNA matrices
• Issues
• License

About TIRmite

TIRmite will use profile-HMM models of Terminal Inverted Repeats (TIRs) for genome-wide annotation of TIR families. These can be provided by the user or built from aligned TIRs oriented as 5' outer edge --> 3' inner edge.

Three classes of output are produced:

1. All significant TIR hit sequences written to fasta (per query HMM).
2. Candidate elements comprised of paired TIRs are written to fasta (per query HMM).
3. Genomic annotations of candidate elements and, optionally, TIR hits (paired and unpaired) are written as a single GFF3 file.

Algorithm overview

1. Use nhmmer genome with TIR-pHMM.
2. Import all hits below --maxeval threshold.
3. For each significant TIR match identify candidate partners, where:
   
   • Is on the same sequence.
     
   • Hit is in complementary orientation.
     
   • Distance is <= --maxdist.
     
   • Hit length is >= model length * --mincov.
     
4. Rank candidate partners by distance downstream of positive-strand hits, and upstream of negative-strand hits.
5. Pair reciprocal top candidate hits.
6. For unpaired hits, find first unpaired candidate partner and check for reciprocity.
7. If the first unpaired candidate is non-reciprocal, check for 2nd-order reciprocity (is outbound top-candidate of current candidate reciprocal.)
8. Iterate steps 6-7 until all TIRs are paired OR number of iterations without new pairing exceeds --stableReps.

Options and usage

Installing TIRmite

TIRmite requires Python >= v3.8

Dependencies:

• TIR-pHMM build and search
  • HMMER3
• Extract terminal repeats from predicted TEs
  • pymummer version >= 0.10.3 with wrapper for nucmer option --diagfactor.
  • MUMmer
  • BLAST+ (Optional)

You can create a Conda environment with these dependencies using the YAML files in this repo.

```bash conda env create -f environment.yml

conda activate tirmite ```

Installation options:

pip install the latest development version directly from this repo.

bash % pip install git+https://github.com/Adamtaranto/TIRmite.git

Install latest release from PyPi.

bash % pip install tirmite

Install from Bioconda.

bash % conda install -c bioconda tirmite

Clone from this repository and install as a local Python package.

Do this if you want to edit the code.

bash git clone https://github.com/Adamtaranto/TIRmite.git && cd TIRmite && pip install -e '.[dev]'

Test installation.

```bash

Print version number and exit.

% tirmite --version tirmite 1.2.0

Get usage information

% tirmite --help ```

Example usage

Report all hits and valid pairings of TIR_A in target.fasta (interval <= 10000, hits cover > 40% len of hmm model), and write GFF3 annotation file.

bash % tirmite --genome target.fasta --hmmFile TIR_A.hmm --gffOut TIR_elements_in_Target.gff3 --maxdist 10000 --mincov 0.4

If you don't have a HMM of your TIR, TIRmite can create one for you using an aligned sample of your TIR with --alnFile.

To skip HMM search and run the pairing algorithm on a custom set of TIR hits (i.e. from blastn), you can provide hits in BED format with --pairbed.

TIRs should always be oriented 5`- 3` with the lefthand TIR.

In this example the two TIRs should be oriented to begin with "GA".

5` GA>>>>>>> ATGC <<<<<<>>>>>>> TACG <<<<<<<AG 5`

Standard options

Run tirmite --help to view the program's most commonly used options:

```code tirmite [-h] [--version] --genome GENOME [--hmmDir HMMDIR] [--hmmFile HMMFILE] [--alnDir ALNDIR] [--alnFile ALNFILE] [--alnFormat {clustal,fasta,nexus,phylip,stockholm}] [--pairbed PAIRBED] [--stableReps STABLEREPS] [--outdir OUTDIR] [--prefix PREFIX] [--nopairing] [--gffOut] [--reportTIR {None,all,paired,unpaired}] [--padlen PADLEN] [--keeptemp] [-v] [--cores CORES] [--maxeval MAXEVAL] [--maxdist MAXDIST] [--nobias] [--matrix MATRIX] [--mincov MINCOV] [--hmmpress HMMPRESS] [--nhmmer NHMMER] [--hmmbuild HMMBUILD]

Info: -h, --help Show this help message and exit --version Show program's version number and exit

Input options: --genome Path to target genome that will be queried with HMMs. Note: Sequence names must be unique. (required) --hmmDir Directory containing pre-prepared TIR-pHMMs. --hmmFile Path to single TIR-pHMM file. Incompatible with "--hmmDir". --alnDir Path to directory containing only TIR alignments to be converted to HMM. --alnFile Provide a single TIR alignment to be converted to HMM. Incompatible with "--alnDir". --alnFormat Alignments provided with "--alnDir" or "--alnFile" are all in this format. Choices=["clustal","fasta","nexus","phylip", "stockholm"] --pairbed If set TIRmite will preform pairing on TIRs from custom bedfile only.

Pairing heuristics: --stableReps Number of times to iterate pairing procedure when no additional pairs are found AND remaining unpaired hits > 0. (Default = 0)

Output and housekeeping: --outdir OUTDIR All output files will be written to this directory. --prefix PREFIX Add prefix to all TIRs and Paired elements detected in this run. Useful when running same TIR-pHMM against many genomes. (Default = None) --nopairing If set, only report TIR-pHMM hits. Do not attempt pairing. (Default = False) --gffOut If set report features as prefix.gff3. File saved to outdir. (Default = False) --reportTIR Options for reporting TIRs in GFF annotation file. Choices=None,'all','paired','unpaired' --padlen Extract x bases either side of TIR when writing TIRs to fasta. (Default = None) --keeptemp If set do not delete temp file directory. (Default = False) -v, --verbose Set syscall reporting to verbose.

HMMER options: --cores Set number of cores available to hmmer software. --maxeval Maximum e-value allowed for valid hit. (Default = 0.001) --maxdist Maximum distance allowed between TIR candidates to consider valid pairing. (Default = None) --nobias Turn OFF bias correction of scores in nhmmer. (Default = False) --matrix Use custom DNA substitution matrix with nhmmer. --mincov Minimum valid hit length as prop of model length. (Default = 0.5)

Non-standard HMMER paths: --hmmpress Set location of hmmpress if not in PATH. --nhmmer Set location of nhmmer if not in PATH. --hmmbuild Set location of hmmbuild if not in PATH. ```

Custom DNA Matrices

nhmmer can be supplied with custom DNA score matrices for assessing hmm match scores. Standard NCBI-BLAST matrices such as NUC.4.4 are compatible. (See: ftp://ftp.ncbi.nlm.nih.gov/blast/matrices/NUC.4.4)

Issues

Submit feedback to the Issue Tracker

License

Software provided under MIT license.