User Guide of SSR2Marker

Author

Junyang Yue

Version

SSR2Marker v1.0

Date

2021-11-28

Introduction

SSR2Marker is an integrated pipeline for identification of SSR markers, classification of SSR categories and design of primer pairs between any two given genome-scale sequences. It can find out both the monomorphic and dimorphic molecular markers with a definite comparison value of sequence length. This program is written in Perl and integrated with BLAST, MAFFT, Primer3 and e-PCR.

• PERL5 is freely available at https://www.perl.org/.
• BLAST is freely available at ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST.
• MAFFT is freely available at https://mafft.cbrc.jp/alignment/software/source.html.
• Primer3 is freely available at https://github.com/primer3-org/primer3.
• e-PCR is freely available at http://ftp.debian.org/debian/pool/main/e/epcr/.

Download

SSR2Marker is freely available at from https://github.com/aaranyue/SSR2Marker.

Install

Users don't need to do anything extra if the dependant softwares are already installed, use it directly !

Usage

perl SSR2Marker.pl [option1] <value1> [option2] <value2> ... [optionN] <valueN>

Options

• -f1 | -fasta1 : A single file in FASTA format containing the genomic or transcriptomic sequence(s) from species A.
• -f2 | -fasta2 : A single file in FASTA format containing the genomic or transcriptomic sequence(s) from species B.
• -m1 | -misa1 : The MISA result of species A produced by the MISA program.
• -m2 | -misa2 : The MISA result of species B produced by the MISA program.
• -m | -motif : Setting the motifs for SSR identification through navigational operations (default: 1=10,2=6,3=5,4=5,5=5,6=5).
• -f | -flanking : Length of the flanking sequences between each SSR locus (must be an integer and larger than 100 (bp), default: 200).
• -v |-version : Showing the version information.
• -h |-help : Showing the help information.

Note : The FASTA files are necessary. The MISA files are optional. If users have existing MISA results, they are suggested to provide them in the current step. This will save about 17% operation times to obtain the SSR primers. While the MISA results were not provided, SSR2Marker will begin with performing the MISA analysis until obtaining the SSR primers. What the users need to know is that the MISA results may present some difference due to the different parameter settings. Anyway, both the FASTA and MISA (if provided) files from two species should be provided together.

Syntax

Syntax 1

Suppose users just have two FASTA files named Hongyang.fasta and White.fasta.

• [user@localhost]$ cd <your directory path>
• [user@localhost]$ perl SSR2Marker.pl -f1 Hongyang.fasta -f2 White.fasta

Syntax 2

Suppose users have two FASTA files as well as their respective MISA results named Hongyang.fasta, Hongyang.fasta.misa, White.fasta and White.fasta.misa.

• [user@localhost]$ cd <your directory path>
• [user@localhost]$ perl SSR2Marker.pl -f1 Hongyang.fasta -f2 White.fasta -m1 Hongyang.fasta.misa -m2 White.fasta.misa

Syntax 3

Suppose users want to set the SSR motifs.

• [user@localhost]$ cd <your directory path>
• [user@localhost]$ perl SSR2Marker.pl -f1 Hongyang.fasta -f2 White.fasta -m

Syntax 4

Suppose users want to set a different length for selection of flanking sequences.

• [user@localhost]$ cd <your directory path>
• [user@localhost]$ perl SSR2Marker.pl -f1 Hongyang.fasta -f2 White.fasta -f 150

Test

The test data could be downloaded from the KGD website: http://kiwifruitgenome.org/, including Hongyang.fasta and White.fasta. Otherwise, users could also use their own sequence data only required in the FASTA format. After obtaining the data and renaming the filenames (e.g., Hongyang.fasta, White.fasta), users are simply needed to type one command to run this pipeline. After running, a total of 10 result files in a new folder, including detailed information of SSR motifs, primer pairs, amplified fragments, sequence sizes, length polymorphisms and statistics calculations, are obtained.

• The genome sequence of Hongyang can be download from the KGD website at http://kiwifruitgenome.org/ftp/A_chinensis/Hongyang/v2.0/Hongyang_genome_v2.0.fa.gz.
• The genome sequence of White can be download from the KGD website at http://kiwifruitgenome.org/ftp/A_eriantha/White/v1.0/White_genome_v1.0.fasta.gz.
• The output files generated by SSR2Marker could be available at https://github.com/aaranyue/SSR2Marker.

Email

Thank you for using SSR2Marker. If you have any question or suggestion, please contact us via the Email address: yuejy@ahau.edu.cn.