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This is the command anfo-tool that can be run in the OnWorks free hosting provider using one of our multiple free online workstations such as Ubuntu Online, Fedora Online, Windows online emulator or MAC OS online emulator

PROGRAM:

NAME


anfo-tool - process native ANFO binary files

SYNOPSIS


anfo-tool [ option | pattern ... ]

DESCRIPTION


anfo-tool is used to filter, process and convert the files created by anfo. Every pattern
on the command line is wildcard expanded, then for every input file (or the standard
input, if no pattern is given), anfo-tool builds a chain of input filters, it then merges
these input streams in one of several ways, splits the result up into multiple output
streams, each of which can have a chain of output filter applied.

OPTIONS


General Options
These options apply globally and modify the behavior of the whole program. They can be
placed anywhere in the command line.

-V, --version
Print version number and exit.

-q, --quiet
Suppress all output except fatal error messages.

-v, --verbose
Produce more output, including progress indicators for most operations.

--debug
Produce debugging output in addition to progress information.

-n, --dry-run
Parse command line, optionally print a description of the intended operations, then
exit.

--vmem X
Limit virtual memory to X megabytes. If memory runs out, anfo-tool tries to free
up memory by forgetting about big files, e.g. genomes. Use this option to avoid
swapping or out-of-memory conditions when operations involve big or multiple
genomes.

Setting Parameters
A parameter can be set multiple times on the command line and will overwrite previous
settings. Any filter option that needs a parameter picks up the last definition that
appeared before the filter option.

--set-slope S
Set the slope parameter to S. The slope is used together with the intercept where
filters apply to alignment scores; alignments scoring no worse than slope * (length
- intercept) are considered good. The default is 7.5.

--set-intercept L
Set the intercept parameter to L. The intercept is used together with the slope
where filters apply to alignment scores; alignments scoring no worse than slope *
(length - intercept) are considered good. The default is 20.

--set-context C
Set the context parameter to C. The context is the number of surrounding bases of
the reference included when printing alignments in text form. The default is 0.

--set-genome G
Set the genome parameter to G. Many filters will only consider the best alignments
to this specific genome if it is set. If no genome is set, the globally best
alignment is used.

--clear-genome
Clear the genome parameter. Filters apply to the globally best alignment
afterwards.

Filter Options
Filters can be applied before merging the inputs or after splitting the back up.

-s, --sort-pos=n
Sort by alignment position while buffering no more than n MiB in memory. If a
genome is set, alignments to that genome are used.

-S, --sort-name=n
Sort by read name while buffering no more than n MiB in memory.

-l, --filter-length=L
Retain alignments only for reads of at least L bases length. The reads themselves
are kept.

-f, --filter-score
Retain alignments only if their score is good enough. Usesslopeandintercept.

--filter-mapq=Q
Remove alignments with mapping quality below Q.

-h, --filter-hit=SEQ
Keep only reads that have a hit to a sequence named SEQ. If SEQ is empty, reads
are kept if they have any hit. If the genome parameter is set, only hits to that
genome count.

--delete-hit=SEQ
Delete alignments to SEQ. If SEQ is empty, all alignments are deleted. If the
genome parameter is set, only alignments to that genome are deleted.

--filter-qual=Q
Mask out bases with quality below Q. Such a base is replaced by the N ambiguity
code.

--multiplicity=N
Keep only reads of molecules that have been sequenced at least N times. Reads are
considered to come from the same original molecule if their aligned coordinates are
identical.

--subsample=F
Subsample a fraction F of the results. Every read is independently and randomly
choosen to be kept or not.

--inside-regions=FILE
Read a list of regions from FILE, then keep only alignments that overlap an
annotated region.

--outside-regions=FILE
Read a list of regions from FILE, then keep only alignments that do not overlap an
annotated region.

Special Filters
-d, --rmdup=Q
Remove PCR duplicates, clamp quality scores to Q. Two reads are considered to be
duplicates, if their aligned coordinates are identical. If a genome is set, the
best alignment to that genome is used, else the globally best alignment. Both
alignments must be good, as determined by slopeandintercept. For a set of
duplicates, a consensus is called, generally increasing the quality scores. If a
resulting quality score exceeds Q, it is set to Q. This filter requires the input
to be sorted by alignment coordinate on the selected genome.

--duct-tape=NAME Duct-tape overlapping alignments into contigs and call a consensus
for them. If a genome is set, alignments to that genome are used, else the
globally best alignments. This filter requires input to be sorted by alignment
coordinate on the genome. Output is a set of contigs, every position gets assigned
a consensus base, a quality score and likelihoods for every possible diallele. (It
is called duct-taping because it kind of looks like an assembly, but is not nearly
as solid.)

--edit-header=ED
Invoke the editor ED on the text representaion of the stream's header. This can be
used to clean up header that have accumulated too much cruft.

Merging Filters
Exactly one merging filter should be given on the command line, all filter options
occuring before that are part of the input filter chains, all further filters become
output chains. If no merging filter is given, --concat is assumed, and all filters are
input filters.

-c, --concat
Concatenate all input streams in the order they appear on the command line.

-m, --merge
Merge sorted input streams, producing a sorted result. All inputs must be sorted
in the same way.

-j, --join
Join input streams and retain the single best hits to each genome. Every input
stream must contain a record for every read, reads are buffered in memory until all
of their hits are collected. This way, joining works well if all inputs are nearly
in the same order. If reads are missing from some streams, joining them will waste
memory.

--mega-merge
Merge many streams such as those produced by running anfo-sge. Streams that
operated on the same reads are joined, then everything is merged.

Output Options
If an output option is given on the command line, the current output filter chain is ended
and a new one is started. If no output option is given, a textual representation of the
final stream is written to stdout. All output options accept - to write to stdout.

-o, --output FILE
Write native binary stream (a compressed protobuf message) to FILE. Writing a
binary stream and reading it back in is lossless.

--output-text FILE
Write protobuf text stream to FILE. If the necessary genomes are available, a
textual representation of the alignments is included. If the context parameter is
set, that many additional bases of the reference upstream and downstream from the
alignment are included.

--output-sam=FILE
Write alignments in SAM format to FILE.

--output-glz FILE
Write contigs in GLZ 0.9 format to FILE. Generating GLZ only works after
application of --duct-tape, every contigs becomes a GLZ record.

--output-3aln FILE
Write contigs in a table based format to FILE. The format is still subject to
change, see the source code for detailed documentation.

--output-fasta FILE
Write alignments(!) in FastA format to FILE. Alignments are writte as pair of
reference and query sequence, aligned coordinates are indicated in the description
of the query sequence. If the context parameter is set, that many additional bases
of the reference upstream and downstream from the alignment are included. This
format is not suggested for any serious use, it exists to support legacy
applications.

--output-fastq FILE
Write sequences(!) in FastQ format to FILE. Writing FastQ effectively
reconstitutes the input to ANFO if no filtering was done on the results.

--output-table FILE
Write per-alignment statistics to FILE. The file has three colums:Âsequence
length, alignment score, difference to next best alignment. It is mainly useful to
analyze/visualize the distribution of alignment scores.

--stats FILE
Write simple statistics to FILE. This results in some simple summary statistics of
a whole stream: number of aligned sequences, average length, GC content.

ENVIRONMENT


ANFO_PATH
Colon separated list of directories searched for genome and index files.

ANFO_TEMP
Temporary space used for sorting of large files.

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