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Result file name — file in SAM format to write the result of the alignment into. This parameter is required.

Library - single-end or paired-end reads.

Prebuilt index — check this box to use an index file instead of a source reference sequence. The index is a set of 6 files with suffixes .1.ebwt, .2.ebwt, .3.ebwt, .4.ebwt, .rev.1.ebwt, and .rev.2.ebwt. The index is created during the alignment. Also you can build it manually.

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Short reads — each added short read is a small DNA sequence file. At least one read should be added.

You can also configure other parameters. They are the same as in the original Bowtie 2 (you can read detailed description of the parameters on the Bowtie 2 manual page).

Select one of the following alignment modes:

The --end-to-end alignment mode:

When the -n mode is selected, Bowtie determines which alignments are valid according to the following policy. Alignments may have no more than N mismatches (where N is a number 0-3) in the first L bases (where L is a number 5 or greater, set with Seed length) on the high-quality (left) end of the read. The sum of the Phred quality values at all mismatched positions (not just in the seed) may not exceed E (set with Maq error). Where qualities are unavailable (e.g. if the reads are from a FASTA file), the Phred quality defaults to 40.

The -v alignment mode:

In -v mode, alignments may have no more than V mismatches, where V may be a number from 0 through 3. Quality values are ignored. The -v mode is mutually exclusive with the -n mode.By default, Bowtie 2 performs end-to-end read alignment. That is, it searches for alignments involving all of the read characters. This is also called an "untrimmed" or "unclipped" alignment.

 When the --local option is specified, Bowtie 2 performs local read alignment. In this mode, Bowtie 2 might "trim" or "clip" some read characters from one or both ends of the alignment if doing so maximizes the alignment score.

The following parameters are available:

Maq error (–maqerr) — maximum permitted total of quality values at all mismatched read positions throughout the entire alignment, not just in the “seed”. The default is 70. By default, Bowtie rounds quality values to the nearest 10 and saturates at 30. Note that the rounding can be disabled with No Maq rounding.

Seed Length (–seedlen) — the number of bases on the high-quality end of the read to which the -n applies. The lowest permitted setting is 5 and the default is 28.

Maximum of backtracks (-maxbts) — the maximum number of backtracks (default: 125 without Best, 800 with Best). A “backtrack” is the introduction of a speculative substitution into the alignment.

Descriptors memory usage (–chunkmbs) — the number of megabytes of memory a given thread is given to store path descriptors in the Best flag. Default: 64. This parameter is available if the Best flag is checkedNumber of mismatches (--N) — sets the number of mismatches to allowed in a seed alignment during multiseed alignment. Can be set to 0 or 1. Setting this higher makes alignment slower (often much slower) but increases sensitivity. 

Seed length (–L) — Sets the length of the seed substrings to align during multiseed alignment. Smaller values make alignment slower but more senstive. 

Add columns to allow gaps (--dpad) — "Pads" dynamic programming problems by <int> columns on either side to allow gaps.

Disallow gaps (–gbar) — disallow gaps within <int> positions of the beginning or end of the read.

Seed (–seed) — use <int> as the seed for pseudo-random number generator.

Threads — launch the specified number of parallel search threads. Threads will run on separate processors/cores and synchronize when parsing reads and outputting alignments.

The following flags are available:

Colorspace (–color) — the input is read in colorspace, colors are encoded as characters A/C/G/T (A=blue, C=green, G=orange, T=red).

No Maq rounding (–nomaqround) — Maq (Mapping and Assembly with Quality) accepts quality values in the Phred quality scale, but internally rounds values to the nearest 10, with a maximum of 30. By default, Bowtie also rounds this way. No Maq rounding prevents this rounding in Bowtie.No unpaired alignments (–no-mixed) — by default, when bowtie2 cannot find a concordant or discordant alignment for a pair, it then tries to find alignments for the individual mates. This option disables that behavior.

No discordant alignments (–no-discordant) — by default, bowtie2 looks for discordant alignments if it cannot find any concordant alignments. A discordant alignment is an alignment where both mates align uniquely, but that does not satisfy the paired-end constraints. This option disables that behavior.

No forward orientation (–nofw) — do if --nofw is specified, bowtie2 will not attempt to align against unpaired reads to the forward (Watson) reference strand. 

No reverse-complement orientation (–norc) — do if --norc is specified, bowtie2 will not attempt to align unpaired reads against the reverse-complement (Crick) reference strand.

Try as hard (–tryhard) — try as hard as possible to find valid alignments when they exist, including paired-end alignments.

Best alignments (–best) — make Bowtie guarantee that reported singleton alignments are “best” in terms of stratum (i.e. number of mismatches, or mismatches in the seed for the case of -n mode) and in terms of the quality values at the mismatched position(s).

All alignments (–all) — report all valid alignments per read or pair. Validity of alignments is determined by the alignment policy (combined effects of -n mode, -v mode, Seed length, and Maq error). 

No overlapping mates (–no-overlap) — if one mate alignment overlaps the other at all, consider that to be non-concordant. 

No mates containing one another (–no-contain) — if one mate alignment contains the other, consider that to be non-concordant. 

Select the required parameters and press the Start button.