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minimap2(1) Bioinformatics tools minimap2(1)

NAME

minimap2 - mapping and alignment between collections of DNA sequences

SYNOPSIS

* Indexing the target sequences (optional):
minimap2 [-x preset] -d target.mmi target.fa
minimap2 [-H] [-k kmer] [-w miniWinSize] [-I batchSize] -d target.mmi target.fa

* Long-read alignment with CIGAR:

minimap2 -a [-x preset] target.mmi query.fa > output.sam
minimap2 -c [-H] [-k kmer] [-w miniWinSize] [...] target.fa query.fa > output.paf

* Long-read overlap without CIGAR:

minimap2 -x ava-ont [-t nThreads] target.fa query.fa > output.paf

DESCRIPTION

Minimap2 is a fast sequence mapping and alignment program that can find overlaps between long noisy reads, or map long reads or their assemblies to a reference genome optionally with detailed alignment (i.e. CIGAR). At present, it works efficiently with query sequences from a few kilobases to ~100 megabases in length at a error rate ~15%. Minimap2 outputs in the PAF or the SAM format.

OPTIONS

Indexing options

-k INT
Minimizer k-mer length [15]
-w INT
Minimizer window size [2/3 of k-mer length]. A minimizer is the smallest k-mer in a window of w consecutive k-mers.
-H
Use homopolymer-compressed (HPC) minimizers. An HPC sequence is constructed by contracting homopolymer runs to a single base. An HPC minimizer is a minimizer on the HPC sequence.
-I NUM
Load at most NUM target bases into RAM for indexing [4G]. If there are more than NUM bases in target.fa, minimap2 needs to read query.fa multiple times to map it against each batch of target sequences. NUM may be ending with k/K/m/M/g/G. NB: mapping quality is incorrect given a multi-part index.
--idx-no-seq
Don't store target sequences in the index. It saves disk space and memory but the index generated with this option will not work with -a or -c. When base-level alignment is not requested, this option is automatically applied.
-d FILE
Save the minimizer index of target.fa to FILE [no dump]. Minimap2 indexing is fast. It can index the human genome in a couple of minutes. If even shorter startup time is desired, use this option to save the index. Indexing options are fixed in the index file. When an index file is provided as the target sequences, options -H, -k, -w, -I will be effectively overridden by the options stored in the index file.

Mapping options

-f FLOAT|INT1[,INT2]
If fraction, ignore top FLOAT fraction of most frequent minimizers [0.0002]. If integer, ignore minimizers occuring more than INT1 times. INT2 is only effective in the --sr or -xsr mode, which sets the threshold for a second round of seeding.
--min-occ-floor INT
Force minimap2 to always use k-mers occurring INT times or less [0]. In effect, the max occurrence threshold is set to the max{INT, -f}.
-g INT
Stop chain enlongation if there are no minimizers within INT-bp [10000].
-r INT
Bandwidth used in chaining and DP-based alignment [500]. This option approximately controls the maximum gap size.
-n INT
Discard chains consisting of <INT number of minimizers [3]
-m INT
Discard chains with chaining score <INT [40]. Chaining score equals the approximate number of matching bases minus a concave gap penalty. It is computed with dynamic programming.
-D
If query sequence name/length are identical to the target name/length, ignore diagonal anchors. This option also reduces DP-based extension along the diagonal.
-P
Retain all chains and don't attempt to set primary chains. Options -p and -N have no effect when this option is in use.
--dual=yes|no
If no, skip query-target pairs wherein the query name is lexicographically greater than the target name [yes]
-X
Equivalent to '-DP --dual=no --no-long-join'. Primarily used for all-vs-all read overlapping.
-p FLOAT
Minimal secondary-to-primary score ratio to output secondary mappings [0.8]. Between two chains overlaping over half of the shorter chain (controlled by -M), the chain with a lower score is secondary to the chain with a higher score. If the ratio of the scores is below FLOAT, the secondary chain will not be outputted or extended with DP alignment later. This option has no effect when -X is applied.
-N INT
Output at most INT secondary alignments [5]. This option has no effect when -X is applied.
-G NUM
Maximum gap on the reference (effective with -xsplice/--splice). This option also changes the chaining and alignment band width to NUM. Increasing this option slows down spliced alignment. [200k]
-F NUM
Maximum fragment length (aka insert size; effective with -xsr/--frag=yes) [800]
-M FLOAT
Mark as secondary a chain that overlaps with a better chain by FLOAT or more of the shorter chain [0.5]
--hard-mask-level
Honor option -M and disable a heurstic to save unmapped subsequences.
--max-chain-skip INT
A heuristics that stops chaining early [50]. Minimap2 uses dynamic programming for chaining. The time complexity is quadratic in the number of seeds. This option makes minimap2 exits the inner loop if it repeatedly sees seeds already on chains. Set INT to a large number to switch off this heurstics.
--no-long-join
Disable the long gap patching heuristic. When this option is applied, the maximum alignment gap is mostly controlled by -r.
--lj-min-ratio FLOAT
Fraction of query sequence length required to bridge a long gap [0.5]. A smaller value helps to recover longer gaps, at the cost of more false gaps.
--splice
Enable the splice alignment mode.
--sr
Enable short-read alignment heuristics. In the short-read mode, minimap2 applies a second round of chaining with a higher minimizer occurrence threshold if no good chain is found. In addition, minimap2 attempts to patch gaps between seeds with ungapped alignment.
--split-prefix STR
Prefix to create temporary files. Typically used for a multi-part index.
--frag=no|yes
Whether to enable the fragment mode [no]
--for-only
Only map to the forward strand of the reference sequences. For paired-end reads in the forward-reverse orientation, the first read is mapped to forward strand of the reference and the second read to the reverse stand.
--rev-only
Only map to the reverse complement strand of the reference sequences.
--heap-sort=no|yes
If yes, sort anchors with heap merge, instead of radix sort. Heap merge is faster for short reads, but slower for long reads. [no]
--no-pairing
Treat two reads in a pair as independent reads. The mate related fields in SAM are still properly populated.

Alignment options

-A INT
Matching score [2]
-B INT
Mismatching penalty [4]
-O INT1[,INT2]
Gap open penalty [4,24]. If INT2 is not specified, it is set to INT1.
-E INT1[,INT2]
Gap extension penalty [2,1]. A gap of length k costs min{O1+k*E1,O2+k*E2}. In the splice mode, the second gap penalties are not used.
-C INT
Cost for a non-canonical GT-AG splicing (effective with --splice) [0]
-z INT1[,INT2]
Truncate an alignment if the running alignment score drops too quickly along the diagonal of the DP matrix (diagonal X-drop, or Z-drop) [400,200]. If the drop of score is above INT2, minimap2 will reverse complement the query in the related region and align again to test small inversions. Minimap2 truncates alignment if there is an inversion or the drop of score is greater than INT1. Decrease INT2 to find small inversions at the cost of performance and false positives. Increase INT1 to improves the contiguity of alignment at the cost of poor alignment in the middle.
-s INT
Minimal peak DP alignment score to output [40]. The peak score is computed from the final CIGAR. It is the score of the max scoring segment in the alignment and may be different from the total alignment score.
-u CHAR
How to find canonical splicing sites GT-AG - f: transcript strand; b: both strands; n: no attempt to match GT-AG [n]
--end-bonus INT
Score bonus when alignment extends to the end of the query sequence [0].
--score-N INT
Score of a mismatch involving ambiguous bases [1].
--splice-flank=yes|no
Assume the next base to a GT donor site tends to be A/G (91% in human and 92% in mouse) and the preceding base to a AG acceptor tends to be C/T [no]. This trend is evolutionarily conservative, all the way to S. cerevisiae (PMID:18688272). Specifying this option generally leads to higher junction accuracy by several percents, so it is applied by default with --splice. However, the SIRV control does not honor this trend (only ~60%). This option reduces accuracy. If you are benchmarking minimap2 on SIRV data, please add --splice-flank=no to the command line.
--end-seed-pen INT
Drop a terminal anchor if s<log(g)+INT, where s is the local alignment score around the anchor and g the length of the terminal gap in the chain. This option is only effective with --splice. It helps to avoid tiny terminal exons. [6]
--no-end-flt
Don't filter seeds towards the ends of chains before performing base-level alignment.
--cap-sw-mem NUM
Skip alignment if the DP matrix size is above NUM. Set 0 to disable [0].

Input/output options

-a
Generate CIGAR and output alignments in the SAM format. Minimap2 outputs in PAF by default.
-Q
Ignore base quality in the input file.
-L
Write CIGAR with >65535 operators at the CG tag. Older tools are unable to convert alignments with >65535 CIGAR ops to BAM. This option makes minimap2 SAM compatible with older tools. Newer tools recognizes this tag and reconstruct the real CIGAR in memory.
-R STR
SAM read group line in a format like @RG\tID:foo\tSM:bar [].
-y
Copy input FASTA/Q comments to output.
-c
Generate CIGAR. In PAF, the CIGAR is written to the `cg' custom tag.
--cs[=STR]
Output the cs tag. STR can be either short or long. If no STR is given, short is assumed. [none]
--MD
Output the MD tag (see the SAM spec).
--eqx
Output =/X CIGAR operators for sequence match/mismatch.
-Y
In SAM output, use soft clipping for supplementary alignments.
--seed INT
Integer seed for randomizing equally best hits. Minimap2 hashes INT and read name when choosing between equally best hits. [11]
-t INT
Number of threads [3]. Minimap2 uses at most three threads when indexing target sequences, and uses up to INT+1 threads when mapping (the extra thread is for I/O, which is frequently idle and takes little CPU time).
-2
Use two I/O threads during mapping. By default, minimap2 uses one I/O thread. When I/O is slow (e.g. piping to gzip, or reading from a slow pipe), the I/O thread may become the bottleneck. Apply this option to use one thread for input and another thread for output, at the cost of increased peak RAM.
-K NUM
Number of bases loaded into memory to process in a mini-batch [500M]. Similar to option -I, K/M/G/k/m/g suffix is accepted. A large NUM helps load balancing in the multi-threading mode, at the cost of increased memory.
--secondary=yes|no
Whether to output secondary alignments [yes]
--max-qlen NUM
Filter out query sequences longer than NUM.
--paf-no-hit
In PAF, output query name and length for an unmapped sequence.
--version
Print version number to stdout

Preset options

-x STR
Preset []. This option applies multiple options at the same time. It should be applied before other options because options applied later will overwrite the values set by -x. Available STR are:
map-pb
PacBio/Oxford Nanopore read to reference mapping (-Hk19)
map-ont
Slightly more sensitive for Oxford Nanopore to reference mapping (-k15). For PacBio reads, HPC minimizers consistently leads to faster performance and more sensitive results in comparison to normal minimizers. For Oxford Nanopore data, normal minimizers are better, though not much. The effectiveness of HPC is determined by the sequencing error mode.
asm5
Long assembly to reference mapping (-k19 -w19 -A1 -B19 -O39,81 -E3,1 -s200 -z200 --min-occ-floor=100). Typically, the alignment will not extend to regions with 5% or higher sequence divergence. Only use this preset if the average divergence is far below 5%.
asm10
Long assembly to reference mapping (-k19 -w19 -A1 -B9 -O16,41 -E2,1 -s200 -z200 --min-occ-floor=100). Up to 10% sequence divergence.
asm20
Long assembly to reference mapping (-k19 -w10 -A1 -B4 -O6,26 -E2,1 -s200 -z200 --min-occ-floor=100). Up to 20% sequence divergence.
ava-pb
PacBio all-vs-all overlap mapping (-Hk19 -Xw5 -m100 -g10000 --max-chain-skip 25).
ava-ont
Oxford Nanopore all-vs-all overlap mapping (-k15 -Xw5 -m100 -g10000 -r2000 --max-chain-skip 25). Similarly, the major difference from ava-pb is that this preset is not using HPC minimizers.
splice
Long-read spliced alignment (-k15 -w5 --splice -g2000 -G200k -A1 -B2 -O2,32 -E1,0 -C9 -z200 -ub --splice-flank=yes). In the splice mode, 1) long deletions are taken as introns and represented as the `N' CIGAR operator; 2) long insertions are disabled; 3) deletion and insertion gap costs are different during chaining; 4) the computation of the `ms' tag ignores introns to demote hits to pseudogenes.
sr
Short single-end reads without splicing (-k21 -w11 --sr --frag=yes -A2 -B8 -O12,32 -E2,1 -r50 -p.5 -N20 -f1000,5000 -n2 -m20 -s40 -g200 -2K50m --heap-sort=yes --secondary=no).

Miscellaneous options

--no-kalloc
Use the libc default allocator instead of the kalloc thread-local allocator. This debugging option is mostly used with Valgrind to detect invalid memory accesses. Minimap2 runs slower with this option, especially in the multi-threading mode.
--print-qname
Print query names to stderr, mostly to see which query is crashing minimap2.
--print-seeds
Print seed positions to stderr, for debugging only.

OUTPUT FORMAT

Minimap2 outputs mapping positions in the Pairwise mApping Format (PAF) by default. PAF is a TAB-delimited text format with each line consisting of at least 12 fields as are described in the following table:
Col Type Description
1 string Query sequence name
2 int Query sequence length
3 int Query start coordinate (0-based)
4 int Query end coordinate (0-based)
5 char `+' if query/target on the same strand; `-' if opposite
6 string Target sequence name
7 int Target sequence length
8 int Target start coordinate on the original strand
9 int Target end coordinate on the original strand
10 int Number of matching bases in the mapping
11 int Number bases, including gaps, in the mapping
12 int Mapping quality (0-255 with 255 for missing)

When alignment is available, column 11 gives the total number of sequence matches, mismatches and gaps in the alignment; column 10 divided by column 11 gives the BLAST-like alignment identity. When alignment is unavailable, these two columns are approximate. PAF may optionally have additional fields in the SAM-like typed key-value format. Minimap2 may output the following tags:

Tag Type Description
tp A Type of aln: P/primary, S/secondary and I,i/inversion
cm i Number of minimizers on the chain
s1 i Chaining score
s2 i Chaining score of the best secondary chain
NM i Total number of mismatches and gaps in the alignment
MD Z To generate the ref sequence in the alignment
AS i DP alignment score
ms i DP score of the max scoring segment in the alignment
nn i Number of ambiguous bases in the alignment
ts A Transcript strand (splice mode only)
cg Z CIGAR string (only in PAF)
cs Z Difference string
dv f Approximate per-base sequence divergence
de f Gap-compressed per-base sequence divergence

The cs tag encodes difference sequences in the short form or the entire query AND reference sequences in the long form. It consists of a series of operations:

Op Regex Description
= [ACGTN]+ Identical sequence (long form)
: [0-9]+ Identical sequence length
* [acgtn][acgtn] Substitution: ref to query
+ [acgtn]+ Insertion to the reference
- [acgtn]+ Deletion from the reference
~ [acgtn]{2}[0-9]+[acgtn]{2} Intron length and splice signal

LIMITATIONS

*
Minimap2 may produce suboptimal alignments through long low-complexity regions where seed positions may be suboptimal. This should not be a big concern because even the optimal alignment may be wrong in such regions.
*
Minimap2 requires SSE2 or NEON instructions to compile. It is possible to add non-SSE2/NEON support, but it would make minimap2 slower by several times.

SEE ALSO

miniasm(1), minimap(1), bwa(1).
10 January 2019 minimap2-2.15 (r905)