Installation
PyroTrimmer was implemented in JAVA, not use any other executable program, so it can be run
on any kind of operating system without requirement for other external programs to be installed.
Prerequsites
To execute PyroTrimmer, You will need the Java Runtime Enviroment(JRE) should be installed on
your computer.
You can get a JRE from http://www.oracle.com/technetwork/java/javase/
downloads/index.html.
Figure A.1
Arguments

Tag: The tag including project, names of genes and samples, length cutoffs of reads, and
sequences of barcodes-linkers-primers. There are two input mode of the tag information; one
is the “from file” and the other “user-typing”.
By clicking of the radio button, you can select this (Figure A.1). If you select “From file” mode,
then the tag information can be imported through file-chooser (Figure A.2).
On the other hand, when you select “User-typing”, you can be imported through file-chooser.
The tag file format must place in order like shown the spreadsheet header (Figure A.2) and be
tab-delaminated list of files with each element (Figure A.3).
Figure A.2
Figure A.3
If the data is single-end, you have to fill the barcode, linker and primer according to strand
and leave blank for other strand. In case of file, that leave blank with tab delimiter. (Figure
A.4a, Figure A.4b)
Blank tab
Figure A.4a (Forward strand)
Blank tab
Figure A.4b (Reverse strand)

Fasta and Quality: The fasta and quality file can be imported using file-chooser.

Output Path:
Sets the name of the directory in which Pyrotrimmer will write all of its output.
The default is the Fasta input path. Output Path can be set using file-chooser.
Options
All option’s value of the PyroTrimmer can be set by the slider control (figure B.1). You can move
slider’s indicator to set value using a mouse easily.

Maximum primer mismatch : Mismatches are allowed to detect priming region (1~4 ,default 3)

3’ end trimming

Window size: Size of window to scan a read from 3 ' to 5 ' direction. Nucleotides are
trimmed out until the moving window from 3’ to 5’ direction meet average quality value
higher than the cutoff value for specified window size. (3~20, default 5)

Average quality value cutoff: Cutoff for trimming low quality 3 ' tails. Nucleotides are
trimmed out until the moving window from 3’ to 5’ direction meet average quality value
higher than the cutoff value for specified window size. (10~20, default 20)

Quality filtering

Maximum ambiguous nucleotide : Removing sequences with more than maximum
ambiguous nucleotide (0~5, default 1)

Average quality value cutoff : Removing Sequences with lower average quality values
than average quality value cutoff for full length of a read (20~30, default25)
Figure B.1
Execution
You can execute the PyroTimmer by clicking “Run” button. When the PyroTrimmer is running, you
can identify process status through number of reads trimmed and progress bar in bottom bar
(Figure C.1).
Figure C.1
After all trimming process are complete, you will show the file tree for the trimmed result on left
side of a frame. (Figure C.2).
Figure C.2
Output
PyroTrimmer generate a number of files that are trimmed result file, summary statics file and
undefined file. (Figure C.2)
Trimmed result files are comprised of Fasta, quality and read length count file.


Fasta and quality: Each Fasta and quality files are named by gene name + sample
number + strand in project directory. A short or long read length files are classified in
“short” or “long” directory separately according to a lower and upper bounds of the tag
information.

Length count file: File that is count per read length in “length_count“ directory.

Result summary file: File in tabular form that contains result count by sample number,
gene, direction, read length, and undefined read.

Undefined Fasta and quality: Reads and quality file which is discarded by options.
Command line version
Arguments :

-t
file path of tag information
-i
file path of the input fasta file
-q
file path of the quality file
-o
file Path for output data
Options :


-a
average quality value cutoff for 3’end trimming: 10 to 30 (default = 20)
-l
average quality value cutoff for full length sequence: 20 to 30 (default = 25)
-m
# of mismatches for trimming primer sequences: 1 to 4 (default = 3)
-p
removing sequences with polynucleotides > arg: 0 to 5 (default = 0)
-w
window size for 3’ end trimming: 3 to 20 (default = 5)
Example :
java -jar PyroTrimmer-1.0.jar -t /home/whoami/tagfile -i /home/whoami/test.fasta -q
/home/whoami/test.quality -o /home/whoami/out -a 20 -l 25 -m 3 -p 0 -w 5