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 [10]. 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 [8G]. 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. This would create a multi-part index. NUM may be ending with k/K/m/M/g/G. NB: mapping quality is incorrect given a multi-part index. See also option --split-prefix.
--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.
--alt FILE: List of ALT contigs [null]
--alt-drop FLOAT: Drop ALT hits by FLOAT fraction when ranking and computing mapping quality [0.15]

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.
-U INT1[,INT2]: Lower and upper bounds of k-mer occurrences [10,1000000]. The final k-mer occurrence threshold is max{INT1, min{INT2, -f}}. This option prevents excessively small or large -f estimated from the input reference. Available since r1034 and deprecating --min-occ-floor in earlier versions of minimap2.
--q-occ-frac FLOAT: Discard a query minimizer if its occurrence is higher than FLOAT fraction of query minimizers and than the reference occurrence threshold [0.01]. Set 0 to disable. Available since r1105.
-e INT: Sample a high-frequency minimizer every INT basepairs [500].
-g NUM: Stop chain enlongation if there are no minimizers within NUM-bp [10k].
-r NUM1[,NUM2]: Bandwidth for chaining and base alignment [500,20k]. NUM1 is used for initial chaining and alignment extension; NUM2 for RMQ-based re-chaining and closing gaps in alignments.
-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]
--rmq=no|yes: Use the minigraph chaining algorithm [no]. The minigraph algorithm is better for aligning contigs through long INDELs.
--rmq-inner NUM: Apply full dynamic programming for anchors within distance NUM [1000].
--hard-mask-level: Honor option -M and disable a heurstic to save unmapped subsequences and disables --mask-len.
--mask-len NUM: Keep an alignment if dropping it leaves an unaligned region on query longer than INT [inf]. Effective without --hard-mask-level.
--max-chain-skip INT: A heuristics that stops chaining early [25]. 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.
--max-chain-iter INT: Check up to INT partial chains during chaining [5000]. This is a heuristic to avoid quadratic time complexity in the worst case.
--chain-gap-scale FLOAT: Scale of gap cost during chaining [1.0]
--no-long-join: Disable the long gap patching heuristic. When this option is applied, the maximum alignment gap is mostly controlled by -r.
--splice: Enable the splice alignment mode.
--sr[=no|dna|rna]: Enable short-read alignment heuristics [no]. If this option is used with no argument, `dna' is set. In the DNA 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-hash-name: Produce the same alignment for identical sequences regardless of their sequence names.

Alignment options

-A INT: Matching score [2]
-B INT: Mismatching penalty [4]
-b INT: Mismatching penalty for transitions [same as -B].
-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.
-J INT: Splice model [1]. 0 for the original minimap2 splice model that always penalizes non-GT-AG splicing; 1 for the miniprot model that considers non-GT-AG. Option -C has no effect with the default -J1.
-j FILE: Junctions used to extend alignment towards ends of reads []. FILE can be gene annotations in the BED12 format (aka 12-column BED), or intron positions in 5-column BED with the strand column required. BED12 file can be converted from GTF/GFF3 with `paftools.js gff2bed anno.gtf'. This option is intended for short RNA-seq reads, while --junc-bed for long noisy RNA-seq reads.
-C INT: Cost for a non-canonical GT-AG splicing (effective with --splice -J0) [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: Penalty of a mismatch involving ambiguous bases [1].
--pairing=strong|weak|no: How to pair paired-end reads [strong]. `no' for aligning the two ends in a pair independently with no `properly paired' set. `weak' for aligning the two ends independently and then pairing the hits. `strong' for jointly aligning and pairing the two ends.
--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.
--spsc FILE: Splice scores []. Each line consists of five fields: 1) contig, 2) offset, 3) `+' or `-', 4) `D' or `A', and 5) score, where offset is the number of bases before a splice junction, `D' indicates the line corresponds to a donor site and `A' for an acceptor site. A positive score suggests the junction is preferred and a negative score suggests the junction is not preferred.
--spsc0 INT: Penalty for positions not in FILE specified by --spsc [5]. Effective with --spsc but not --junc-bed.
--spsc-scale FLOAT: Scale splice scores in --spsc by FLOAT rounded to the nearest integer [0.7].
--junc-bed FILE: Junctions to prefer during base alignment []. Same format as -j. It is NOT recommended to apply this option to short RNA-seq reads. This would increase run time with little improvement to junction accuracy.
--junc-bonus INT: Score bonus for a splice donor or acceptor found in annotation [9]. Effective with --junc-bed but not --spsc.
--jump-min-match INT: Minimum matching length to create a jump [3]. Equivalent to STAR --alignSJDBoverhangMin.
--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 [100m].
--cap-kalloc NUM: Free thread-local kalloc memory reservoir if after the alignment the size of the reservoir above NUM. Set 0 to disable [500m].

Input/output options

-a: Generate CIGAR and output alignments in the SAM format. Minimap2 outputs in PAF by default.
-o FILE: Output alignments to FILE [stdout].
-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[=short|long]: Output the cs tag. If no argument is given, `short' is set. [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.
--secondary-seq: In SAM output, show query sequences for secondary alignments.
--write-junc: Output splice junctions in 6-column BED: contig name, start, end, read name, score and strand. Score is the sum of donor and acceptor scores, where GT gets 3, GC gets 2 and AT gets 1 at donor sites, while AG gets 3 and AC gets 1 at acceptor sites. Alignments with mapping quality below 10 are ignored.
--pass1 FILE: Junctions BED file outputted by --write-junc []. Rows with scores lower than 5 are ignored. When both -j and --pass1 are present, junctions in -j are preferred over in --pass1 when there is ambiguity.
--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 unmapped queries; the strand and the reference name fields are set to `*'. Warning: some paftools.js commands may not work with such output for the moment.
--sam-hit-only: In SAM, don't output unmapped reads.
--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-ont: Align noisy long reads of ~10% error rate to a reference genome. This is the default mode.
lr:hq: Align accurate long reads (error rate <1%) to a reference genome (-k19 -w19 -U50,500 -g10k). This was recommended by ONT developers for recent Nanopore reads produced with chemistry v14 that can reach ~99% in accuracy. It was shown to work better for accurate Nanopore reads than map-hifi.
map-hifi: Align PacBio high-fidelity (HiFi) reads to a reference genome (-xlr:hq -A1 -B4 -O6,26 -E2,1 -s200). It differs from lr:hq only in scoring. It has not been tested whether lr:hq would work better for PacBio HiFi reads.
map-pb: Align older PacBio continuous long (CLR) reads to a reference genome (-Hk19). Note that this data type is effectively deprecated by HiFi. Unless you work on very old data, you probably want to use map-hifi or lr:hq.
map-iclr: Align Illumina Complete Long Reads (ICLR) to a reference genome (-k19 -B6 -b4 -O10,50). This was recommended by Illumina developers.
asm5: Long assembly to reference mapping (-k19 -w19 -U50,500 --rmq -r1k,100k -g10k -A1 -B19 -O39,81 -E3,1 -s200 -z200 -N50). Typically, the alignment will not extend to regions with 5% or higher sequence divergence. Use this preset if the average divergence is not much higher than 0.1%.
asm10: Long assembly to reference mapping (-k19 -w19 -U50,500 --rmq -r1k,100k -g10k -A1 -B9 -O16,41 -E2,1 -s200 -z200 -N50). Use this if the average divergence is around 1%.
asm20: Long assembly to reference mapping (-k19 -w10 -U50,500 --rmq -r1k,100k -g10k -A1 -B4 -O6,26 -E2,1 -s200 -z200 -N50). Use this if the average divergence is around several percent.
splice: Long-read spliced alignment (-k15 -w5 --splice -g2k -G200k -A1 -B2 -O2,32 -E1,0 -C9 -z200 -ub --junc-bonus=9 --cap-sw-mem=0 --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.
splice:hq: Spliced alignment for accurate long RNA-seq reads such as PacBio iso-seq (-xsplice -C5 -O6,24 -B4).
splice:sr: Spliced alignment for short RNA-seq reads (-xsplice:hq --frag=yes -m25 -s40 -2K100m --heap-sort=yes --pairing=weak --sr=rna --min-dp-len=20 --secondary=no).
sr: Short-read alignment without splicing (-k21 -w11 --sr --frag=yes -A2 -B8 -O12,32 -E2,1 -r100 -p.5 -N20 -f1000,5000 -n2 -m25 -s40 -g100 -2K50m --heap-sort=yes --secondary=no).
ava-pb: PacBio CLR all-vs-all overlap mapping (-Hk19 -Xw5 -e0 -m100).
ava-ont: Oxford Nanopore all-vs-all overlap mapping (-k15 -Xw5 -e0 -m100 -r2k).

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
SA	Z	List of other supplementary alignments (with approximate CIGAR strings)
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
rl	i	Length of query regions harboring repetitive seeds
zd	i	Alignment broken due to Z-drop; bit 1: left broken; bit 2: right broken

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.