Primer size (bp)
Maximum number of nucleotides in the primer
Default: 20
Primer overlap length (bp)
Number of bases primers can overlap by in design phase
Default: 19
Minimum GC percentage
Minimum GC content of each primer
Default: 45
Maximum GC percentage
Maximum GC content of each primer
Default: 55
Minimum Tm (°C)
Lower limit of primer melting temperatures
Default: 56
Maximum Tm (°C)
Upper limit of primer melting temperatures
Default: 65
Maximum number of degenerate bases allowed (bp)
Maximum number of IUPAC bases allowed in final primer
Default: 3
Maximum number of monomer bases allowed (bp)
Maximum Number of bases in a mononucleotide run within a primer
Default: 4
Primer concentration (nM)
Concentration at which primer will be synthesized (used for Tm calculation)
Default: 50
Salt concentration (mM)
Salt concentration in primer stock (used for Tm calculation)
Default: 50
Valid base minimum frequency
The minimum frequency of a given base at a given position in the alignment at which the base is considered valid
Default: 0.1
Valid degenerate maximum frequency
The upper limit of conservation in an alignment at a given position, above which the base is called for the majority at that position. Below this, a dengenerate base is called.
Default: 0.85
Maximum self-complement bases allowed within a single primer (bp)
Maximum number of bases within a primer that are self-complimentary
Default: 10
Valid gap minimum frequency
The frequency of genomes within an alignment that must contain a gap at a given position for that gap to be represented in the consensus
Default: 0.8
Primer blast minimum hit frequency
Minimum frequency of hits for a given primer back to the genomes in the alignment
Default: 0.8
Maximum amplicon length (bp)
Upper limit for amplified product length
Default: 900
Minimum amplicon length (bp)
Lower limit for amplified product length
Default: 500
Amplicon overlap length (bp)
Desired overlap between adjacent amplicons in the panel
Default: 100
Maximum degenerate primer combination
Maximum number of unique primer combinations as determined by the permutations of degenerate bases within a primer
Default: 8
Maximum complement bases in primer pair
Maximum number of bases that can be complimentary between a forward and reverse primer in a pair
Default: 15
Genome name
Single word (no spaces or symbols) to indicate template. This title will appear in primer name.
Upload genome multi-fasta file (required)
The genome multi-fasta file contains the sequence of all genomes of your project in fasta format. The genome fasta file is the input file when you use MUSCLE or CLUSTALW to generate alignment file. No space in file name. For example, HIV_AE.fasta
Upload alignment file (Optional)
The multiple alignment file is the output of MUSCLE, or CLUSTALW. PriSM only accepts the output from MUSCLE and CLUSTALW. The file should be named with file extension ".muscle" or ".aln". No space in file name. For example, HIV_AE.muscle, HIV_C.aln
If user does not upload alignment file, PriSM will use MUSCLE to generate alignment using MUSCLE default conditions.
Copyright © Special Project Group, Broad Institute of MIT and Harvard Last Modified 06-05-2009