A wrapper for guppy basecaller. This expects two type of inputs: a collection of fast5 files, and a configuration in the form of a tar file.
You can find configurations at https://github.com/nanoporetech/rerio, and in particular the directory https://github.com/nanoporetech/rerio/basecall_models.
Each file there contains a URL you can download to use, for example https://github.com/nanoporetech/rerio/blob/master/basecall_models/res_rna2_r941_min_flipflop_v001 points to 'https://nanoporetech.box.com/shared/static/84e1jeudx8lr8ay7e9u1ebnvx3bk2kjf.tgz'
When uploading these .tgz files take care to set the format to 'tar' (galaxy doesn't autodetect this?).
The results should be fastq files.
guppy_basecaller --help : Guppy Basecalling Software, (C) Oxford Nanopore Technologies, Limited. Version 3.6.1+249406c, client-server API version 1.1.0
Usage:
With config file:
guppy_basecaller -i <input path> -s <save path> -c <config file> [options]
With flowcell and kit name:
guppy_basecaller -i <input path> -s <save path> --flowcell <flowcell name> --kit <kit name>
List supported flowcells and kits:
guppy_basecaller --print_workflows
Use server for basecalling:
guppy_basecaller -i <input path> -s <save path> -c <config file> --port <server address> [options]
Command line parameters:
--trim_threshold arg Threshold above which data will be trimmed
(in standard deviations of current level
distribution).
--trim_min_events arg Adapter trimmer minimum stride intervals
after stall that must be seen.
--max_search_len arg Maximum number of samples to search through
for the stall
--override_scaling Manually provide scaling parameters rather
than estimating them from each read.
--scaling_med arg Median current value to use for manual
scaling.
--scaling_mad arg Median absolute deviation to use for manual
scaling.
--trim_strategy arg Trimming strategy to apply: 'dna' or 'rna'
(or 'none' to disable trimming)
--dmean_win_size arg Window size for coarse stall event
detection
--dmean_threshold arg Threhold for coarse stall event detection
--jump_threshold arg Threshold level for rna stall detection
--pt_scaling Enable polyT/adapter max detection for read
scaling.
--pt_median_offset arg Set polyT median offset for setting read
scaling median (default 2.5)
--adapter_pt_range_scale arg Set polyT/adapter range scale for setting
read scaling median absolute deviation
(default 5.2)
--pt_required_adapter_drop arg Set minimum required current drop from
adapter max to polyT detection. (default
30.0)
--pt_minimum_read_start_index arg Set minimum index for read start sample
required to attempt polyT scaling. (default
30)
--as_model_file arg Path to JSON model file for adapter
scaling.
--as_gpu_runners_per_device arg Number of runners per GPU device for
adapter scaling.
--as_cpu_threads_per_scaler arg Number of CPU worker threads per adapter
scaler
--as_reads_per_runner arg Maximum reads per runner for adapter
scaling.
--as_num_scalers arg Number of parallel scalers for adapter
scaling.
-m [ --model_file ] arg Path to JSON model file.
-k [ --kernel_path ] arg Path to GPU kernel files location (only
needed if builtin_scripts is false).
-x [ --device ] arg Specify basecalling device: 'auto', or
'cuda:<device_id>'.
--builtin_scripts arg Whether to use GPU kernels that were
included at compile-time.
--chunk_size arg Stride intervals per chunk.
--chunks_per_runner arg Maximum chunks per runner.
--chunks_per_caller arg Soft limit on number of chunks in each
caller's queue. New reads will not be
queued while this is exceeded.
--high_priority_threshold arg Number of high priority chunks to process
for each medium priority chunk.
--medium_priority_threshold arg Number of medium priority chunks to process
for each low priority chunk.
--overlap arg Overlap between chunks (in stride
intervals).
--gpu_runners_per_device arg Number of runners per GPU device.
--cpu_threads_per_caller arg Number of CPU worker threads per
basecaller.
--num_callers arg Number of parallel basecallers to create.
--post_out Return full posterior matrix in output
fast5 file and/or called read message from
server.
--stay_penalty arg Scaling factor to apply to stay probability
calculation during transducer decode.
--qscore_offset arg Qscore calibration offset.
--qscore_scale arg Qscore calibration scale factor.
--temp_weight arg Temperature adjustment for weight matrix in
softmax layer of RNN.
--temp_bias arg Temperature adjustment for bias vector in
softmax layer of RNN.
--qscore_filtering Enable filtering of reads into PASS/FAIL
folders based on min qscore.
--min_qscore arg Minimum acceptable qscore for a read to be
filtered into the PASS folder
--reverse_sequence arg Reverse the called sequence (for RNA
sequencing).
--u_substitution arg Substitute 'U' for 'T' in the called
sequence (for RNA sequencing).
--log_speed_frequency arg How often to print out basecalling speed.
--barcode_kits arg Space separated list of barcoding kit(s) or
expansion kit(s) to detect against. Must be
in double quotes.
--trim_barcodes Trim the barcodes from the output sequences
in the FastQ files.
--num_extra_bases_trim arg How vigorous to be in trimming the barcode.
Default is 0 i.e. the length of the
detected barcode. A positive integer means
extra bases will be trimmed, a negative
number is how many fewer bases (less
vigorous) will be trimmed.
--arrangements_files arg Files containing arrangements.
--score_matrix_filename arg File containing mismatch score matrix.
--start_gap1 arg Gap penalty for aligning before the
reference.
--end_gap1 arg Gap penalty for aligning after the
reference.
--open_gap1 arg Penalty for opening a new gap in the
reference.
--extend_gap1 arg Penalty for extending a gap in the
reference.
--start_gap2 arg Gap penalty for aligning before the query.
--end_gap2 arg Gap penalty for aligning after the query.
--open_gap2 arg Penalty for opening a new gap in the query.
--extend_gap2 arg Penalty for extending a gap in the query.
--min_score arg Minimum score to consider a valid
alignment.
--min_score_rear_override arg Minimum score to consider a valid alignment
for the rear barcode only (and min_score
will then be used for the front only when
this is set).
--front_window_size arg Window size for the beginning barcode.
--rear_window_size arg Window size for the ending barcode.
--require_barcodes_both_ends Reads will only be classified if there is a
barcode above the min_score at both ends of
the read.
--allow_inferior_barcodes Reads will still be classified even if both
the barcodes at the front and rear (if
applicable) were not the best scoring
barcodes above the min_score.
--detect_mid_strand_barcodes Search for barcodes through the entire
length of the read.
--min_score_mid_barcodes arg Minimum score for a barcode to be detected
in the middle of a read.
--num_barcoding_buffers arg Number of GPU memory buffers to allocate to
perform barcoding into. Controls level of
parallelism on GPU for barcoding.
--num_barcode_threads arg Number of worker threads to use for
barcoding.
--calib_detect Enable calibration strand detection and
filtering.
--calib_reference arg Reference FASTA file containing calibration
strand.
--calib_min_sequence_length arg Minimum sequence length for reads to be
considered candidate calibration strands.
--calib_max_sequence_length arg Maximum sequence length for reads to be
considered candidate calibration strands.
--calib_min_coverage arg Minimum reference coverage to pass
calibration strand detection.
--print_workflows Output available workflows.
--flowcell arg Flowcell to find a configuration for
--kit arg Kit to find a configuration for
-z [ --quiet ] Quiet mode. Nothing will be output to
STDOUT if this option is set.
--trace_categories_logs arg Enable trace logs - list of strings with
the desired names.
--verbose_logs Enable verbose logs.
--disable_pings Disable the transmission of telemetry
pings.
--ping_url arg URL to send pings to
--ping_segment_duration arg Duration in minutes of each ping segment.
-q [ --records_per_fastq ] arg Maximum number of records per fastq file, 0
means use a single file (per worker, per
run id).
--read_batch_size arg Maximum batch size, in reads, for grouping
input files.
--compress_fastq Compress fastq output files with gzip.
-i [ --input_path ] arg Path to input fast5 files.
--input_file_list arg Optional file containing list of input
fast5 files to process from the input_path.
-s [ --save_path ] arg Path to save fastq files.
-l [ --read_id_list ] arg File containing list of read ids to filter
to
-r [ --recursive ] Search for input files recursively.
--fast5_out Choice of whether to do fast5 output.
--resume Resume a previous basecall run using the
same output folder.
--progress_stats_frequency arg Frequency in seconds in which to report
progress statistics, if supplied will
replace the default progress display.
--max_block_size arg Maximum block size (in events) of basecall
messages to server.
-p [ --port ] arg Port for basecalling service.
--barcoding_config_file arg Configuration file to use for barcoding.
--num_barcode_threads arg Number of worker threads to use for
barcoding.
--disable_events Disable the transmission of event tables
when receiving reads back from the basecall
server.
--client_id arg Optional unique identifier (non-negative
integer) for this instance of the Guppy
Client Basecaller, if supplied will form
part of the output filenames.
--nested_output_folder If flagged output fastq files will be
written to a nested folder structure, based
on: protocol_group/sample/protocol/qscore_p
ass_fail/barcode_arrangement/
-h [ --help ] produce help message
-v [ --version ] print version number
-c [ --config ] arg Config file to use
-d [ --data_path ] arg Path to use for loading any data files the
application requires.