tirmite 1.1.1

Map TIR-pHMM models to genomic sequences for annotation of MITES and complete DNA-Transposons.

# 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.


TIRmite is packaged with *tSplit* a tool for extraction of terminal repeats
from complete transposons.

# Table of contents

* [About TIRmite](#about-tirmite)
* [Algorithm overview](#algorithm-overview)
* [Options and usage](#options-and-usage)
* [Installing TIRmite](#installing-tirmite)
* [Example usage](#example-usage)
* [Standard options](#standard-options)
* [Custom DNA matrices](#custom-dna-matrices)
* [Additional tools](additional-tools)
* [tSplit](tsplit)
* [tSplit algorithm overview](tsplit-algorithm-overview)
* [tSplit options and usage](tsplit-options-and-usage)
* [tSplit example usage](tsplit-example-usage)
* [tsplit-LTR](tsplit-ltr)
* [tsplit-TIR](tsplit-tir)
* [tSplit options](tsplit-options)
* [License](#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*
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

Dependencies:
- TIR-pHMM build and search
* [HMMER3](http://hmmer.org)
- Optional for experimental Bowtie2 mapping mode:
- [bowtie2](http://bowtie-bio.sourceforge.net/bowtie2/index.shtml)
- [samtools](https://github.com/samtools/samtools)
- [bedtools](http://bedtools.readthedocs.io/en/latest/)
- Extract terminal repeats from predicted TEs
* [pymummer](https://pypi.python.org/pypi/pymummer) version >= 0.10.3 with wrapper for nucmer option *--diagfactor*.
* [MUMmer](http://mummer.sourceforge.net/)
* [BLAST+](ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST/) (Optional)

Installation options:

```bash
# Install from PyPi:
pip install tirmite

# Clone and install from this repository:
git clone https://github.com/Adamtaranto/TIRmite.git && cd TIRmite && pip install -e .
```

## Example usage

Report all hits and valid pairings of TIR_A in target.fasta (interval <= 50000),
and write GFF3 annotation file.

```
tirmite --genome target.fasta --hmmFile TIR_A.hmm --gffOut TIR_elements_in_Target.gff3 --maxdist 50000
```

## Standard options

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

```
Usage: tirmite [-h] --genome GENOME [--hmmDir HMMDIR] [--hmmFile HMMFILE]
[--alnDir ALNDIR] [--alnFile ALNFILE]
[--alnFormat {clustal,emboss,fasta,fasta-m10,ig,maf,mauve,nexus,phylip,phylip-sequential,phylip-relaxed,stockholm}]
[--useBowtie2] [--btTIR BTTIR] [--bowtie2 BOWTIE2]
[--bt2build BT2BUILD] [--samtools SAMTOOLS]
[--bedtools BEDTOOLS] [--stableReps STABLEREPS]
[--outdir OUTDIR] [--prefix PREFIX] [--nopairing]
[--gffOut GFFOUT] [--reportTIR {None,all,paired,unpaired}]
[--keeptemp] [-v] [--cores CORES] [--maxeval MAXEVAL]
[--maxdist MAXDIST] [--nobias] [--matrix MATRIX]
[--hmmpress HMMPRESS] [--nhmmer NHMMER] [--hmmbuild HMMBUILD]

Help:
-h, --help Show this help message 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 the 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","emboss","fasta","fasta-m10","ig","maf","mauve",
"nexus","phylip","phylip-sequential","phylip-relaxed","stockholm"]


Alternative search methods:
--useBowtie2 If set, map short TIR to genome with bowtie2.
Experimental method, potentially useful for very short though highly conserved TIRs where
TIR-pHMM hits return high e-values.
--btTIR Fasta file containing a single TIR to be mapped with bowtie2.
--bowtie2 Set location of bowtie2 if not in PATH.
--bt2build Set location of bowtie2-build if not in PATH.
--samtools Set location of samtools if not in PATH.
--bedtools Set location of bedtools if not in PATH.


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 All output files will be written to this directory.
--gffOut GFF3 annotation filename.
--reportTIR Options for reporting TIRs in GFF annotation file.
Choices=[None,'all','paired','unpaired']
(Default = 'all')
--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)
--keeptemp If set do not delete temp file directory.
(Default = False)
-v, --verbose Set syscall reporting to verbose.

HMMER options:
--cores Set the number of cores available to hmmer software.
(Default = 1)
--maxeval Maximum e-value allowed for valid hit.
(Default = 0.001)
--maxdist Maximum distance allowed between TIR candidates to consider valid pairing.
--nobias Turn OFF bias correction of scores in nhmmer.
(Default = False)
--matrix Use custom DNA substitution matrix with nhmmer.


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)

# Additional tools

# tSplit

Extract terminal repeats from retrotransposons (LTRs) or DNA transposons (TIRs).

# tSplit algorithm overview

tSplit attempts to identify terminal repeats in transposable elements by
first aligning each element to itself using nucmer, and then applying a set of
tuneable heuristics to select an alignment pair most likely to represent an LTR or TIR.

1. Exclude all diagonal/self-matches
2. If tsplit-LTR: Retain only alignment pairs on the same strand (tandem repeats)
3. If tsplit-TIR: Retain only alignment pairs on opposite strands (inverse repeats)
4. Retain pairs for which the 5' match begins within x bases of element start
and whose 3' match ends within x bases of element end
5. Exclude alignment pairs which overlap (potential SSRs)
6. If multiple candidates remain select alignment pair with largest internal segment
(i.e. closest to element ends)

# tSplit options and usage

## tSplit example usage

TE-splitter contains two programs: tsplit-LTR and tsplit-TIR, for extracting long terminal
repeats and terminal inverted repeats, respectively. Options are the same
for each.

## tsplit-LTR

For each element in *retroelements.fasta* split into internal and external segments.
Split segments will be written to *LTR_split_TE-splitter_output.fasta* with suffix "_I"
for internal or "_LTR" for external segments. LTRs must be at least 10bp in length and
share 80% identity and occur within 10bp of each end of the input element.

```bash
tsplit-LTR -i retroelements.fasta -p LTR_split
```

## tsplit-TIR

For each element in *dna-transposons.fasta* split into internal and external (TIR) segments.
Split segments will be written to *TIR_split_TE-splitter_output.fasta* with suffix "_I" for
internal or "_TIR" for external segments. TIRs must be at least 10bp in length and share 80%
identity and occur within 10bp of each end of the input element. Additionally, synthetic
MITEs will be constructed by concatenation of left and right TIRs, with internal segments
excised.

```bash
tsplit-TIR -i dna-transposons.fasta -p TIR_split --makemites
```

## tSplit options

Run `tsplit-LTR --help` or `tsplit-TIR --help` to view the programs' most commonly used
options:

```
Usage: tsplit-[LTR or TIR] [-h] -i INFILE [-p PREFIX] [-d OUTDIR]
[--splitmode {all,split,internal,external,None}]
[--makemites] [--keeptemp] [-v] [-m MAXDIST]
[--minid MINID] [--minterm MINTERM] [--minseed MINSEED]
[--diagfactor DIAGFACTOR] [--method {blastn,nucmer}]

Help:
-h, --help Show this help message and exit.

Input:
-i, --infile Multifasta containing complete elements.
(Required)

Output:
-p, --prefix All output files begin with this string. (Default:[infile basename])
-d, --outdir Write output files to this directory. (Default: cwd)
--keeptemp If set do not remove temp directory on completion.
-v, --verbose If set, report progress.

Report settings:
--splitmode Options: {all,split,internal,external,None}
all = Report input sequence as well as internal and external segments.
split = Report internal and external segments after splitting.
internal = Report only internal segments.
external = Report only terminal repeat segments.
None = Only report synthetic MITES (when --makemites is also set).
(Default: split)
--makemites Experimental function: Attempt to construct synthetic MITE sequences from TIRs by concatenating
5' and 3' TIRs. Available only in 'tsplit-TIR' mode

Alignment settings:
--method Select alignment tool. Note: blastn may perform better on very short high-identity TRs,
while nucmer is more robust to small indels.
Options: {blastn,nucmer}
(Default: nucmer)
--minid Minimum identity between terminal repeat pairs. As float.
(Default: 80.0)
--minterm Minimum length for a terminal repeat to be considered.
Equivalent to nucmer "--mincluster"
(Default: 10)
-m, --maxdist Terminal repeat candidates must be no more than this many bases from ends of an input element.
Note: Increase this value if you suspect that your element is nested within some flanking sequence.
(Default: 10)
--minseed Minimum length of a maximal exact match to be included in final match cluster.
Equivalent to nucmer "--minmatch".
(Default: 5)
--diagfactor Maximum diagonal difference factor for clustering of matches within nucmer,
i.e. diagonal difference / match separation
(default 0.20)
Note: Increase value for greater tolerance of indels between terminal repeats.
```

# License

Software provided under MIT license.