PCRpanel

Custom Amplicon Panels Designer is a professional tool for amplicon sequencing technology, an ultra-high multiplex tiling PCR sequencing approach targeting DNA sequencing technology for Next-Generation Sequencing (NGS) and Nanopore sequencing (ONT).

The scope of this application is not limited to multiplex tiling PCR. Any sequence, of any length, and any number of target sequences can be used. There are no restrictions on the size of amplicons or their number, and standard PCR tasks for multiplex applications can be performed. In the target sequence, only the coordinates for analysis need to be specified; these can be exons, introns, or absolutely any task. The user can develop PCR sets based on an existing list of primers or probes from previously developed panels or for other purposes. Non-specific tails for primers can be added during the primer design stage.

By Ruslan Kalendar

email: [email protected]

Online version

Availability and requirements:

Operating system(s): Platform independent

Programming language: Java 24 or higher

Java Downloads

How do I set or change the Java path system variable

To run the project from the command line. Command-line options, separated by spaces. The executive file PCRpanel.jar is in the dist directory, which can be copied to any location. Go to the target folder and type the following; an individual file or a file folder can be specified:

java -jar <PCRpanelPath>\dist\PCRpanel.jar <PCRpanelPath>\test\config.file

java -jar C:\PCRpanel\dist\PCRpanel.jar C:\PCRpanel\test\config.file 

Basic usage:

To enter parameters and specify the location of the target files and primer's file, you must specify this via a file on the command line. An example of such a file here (file name or extension does not matter):

config.file

target_path=C:\PCRpanel\test\NG_008690.txt
target_path=C:\PCRpanel\test\NG_011731.txt
target_path=C:\PCRpanel\test\NG_013019.txt
target_path=C:\PCRpanel\test\NG_008847.txt
target_path=C:\PCRpanel\test\NC_000002.txt

target_primers=C:\PCRpanel\test\primers.txt

minPCR=250
maxPCR=500
minLen=18
maxLen=24
minTm=60
maxTm=62
minLC=78
3end=w
5end=
forwardtail=ACACTCTTTCCCTACACGACGCTCTTCCGATCT
reversetail=GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT

Specifies the coordinates of exons to be analyzed:

In the Genbank file ("FEATURES"), you must replace "mRNA" with "Panel" to indicate exon coordinates or other target fragments to the software.

Example: Homo sapiens HNF1 homeobox A: https://www.ncbi.nlm.nih.gov/nuccore/NG_011731.2?from=4823&to=28767&report=genbank

 mRNA            join(179..527,10266..10465,14953..15139,15597..15838,
                 17695..17846,17974..18175,18907..19098,20701..20822,
                 20916..21060,22498..23945)

replaced by:

 Panel           join(179..527,10266..10465,14953..15139,15597..15838,
                 17695..17846,17974..18175,18907..19098,20701..20822,
                 20916..21060,22498..23945)

Specifies the coordinates for developing a panel for the entire length:

Example: Severe acute respiratory syndrome coronavirus 2: https://www.ncbi.nlm.nih.gov/nuccore/NC_045512.2

For whole-genome tiling, it is necessary to specify the coordinates for developing a panel for the entire length of the virus genome.

Insert the following line ("Panel join(1..29842)") under the ("FEATURES") in this the Genbank file:

FEATURES             Location/Qualifiers
     Panel           join(1..29842)

The target sequence is not limited by the presence of exons, as in the example with the HNF1 gene. Anyone can specify any coordinates and any number of them for any target sequence.