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eprimer3.yaml
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!mobyle/program
name: eprimer3
title: eprimer3
description: Picks PCR primers and hybridization oligos
inputs: !mobyle/inputparagraph
children:
- !mobyle/inputprogramparagraph
prompt: Input section
name: input
children:
- !mobyle/inputprogramparameter
comment: The sequence from which to choose primers. The sequence must
be presented 5' to 3'
prompt: Sequence option
format: ("", " -sequence=" + str(value))[value is not None]
simple: true
argpos: 1
mandatory: true
name: sequence
command: false
type: !mobyle/formattedtype
format_terms: ['EDAM_format:1927', 'EDAM_format:2200', 'EDAM_format:1935',
'EDAM_format:1936', 'EDAM_format:1948', 'EDAM_format:1948', 'EDAM_format:1957',
'EDAM_format:2188']
data_terms: ['EDAM_data:2977']
- !mobyle/inputprogramparameter
comment: Tell EPrimer3 to pick primer(s)
prompt: Pick pcr primer(s)
format: (" -noprimer", "")[ bool(value) ]
argpos: 2
name: primer
command: false
type: !mobyle/booleantype {default: false}
- !mobyle/inputprogramparameter
comment: 'Tell EPrimer3 what task to perform. Legal values are 1: ''Pick
PCR primers'', 2: ''Pick forward primer only'', 3: ''Pick reverse
primer only'', 4: ''No primers needed''.'
prompt: Task
format: ("", " -task=" + str(value))[value is not None and value!=vdef]
argpos: 3
name: task
precond: primer
command: false
type: !mobyle/stringtype
default: '1'
options:
- {label: Pick pcr primers, value: '1'}
- {label: Pick forward primer only, value: '2'}
- {label: Pick reverse primer only, value: '3'}
- {label: No primers needed, value: '4'}
- !mobyle/inputprogramparameter
comment: An 'internal oligo' is intended to be used as a hybridization
probe (hyb probe) to detect the PCR product after amplification.
prompt: Pick hybridization probe
format: ("", " -hybridprobe")[ bool(value) ]
argpos: 4
name: hybridprobe
command: false
type: !mobyle/booleantype {default: false}
- !mobyle/inputprogramparameter {comment: "Similar to MISPRIMING-LIBRARY,\
\ except that the event we seek to avoid is hybridization of the internal\
\ oligo to sequences in this library rather than priming from them.\
\ \n The file must be in (a slightly restricted) FASTA format (W.\
\ B. Pearson and D.J. Lipman, PNAS 85:8 pp 2444-2448 [1988]); we\
\ briefly discuss the organization of this file below. If this parameter\
\ is specified then EPrimer3 locally aligns each candidate oligo\
\ against each library sequence and rejects those primers for which\
\ the local alignment score times a specified weight (see below)\
\ exceeds INTERNAL-OLIGO-MAX-MISHYB. (The maximum value of the weight\
\ is arbitrarily set to 12.0.) \n Each sequence entry in the FASTA-format\
\ file must begin with an 'id line' that starts with '>'. The contents\
\ of the id line is 'slightly restricted' in that EPrimer3 parses\
\ everything after any optional asterisk ('*') as a floating point\
\ number to use as the weight mentioned above. If the id line contains\
\ no asterisk then the weight defaults to 1.0. The alignment scoring\
\ system used is the same as for calculating complementarity among\
\ oligos (e.g. SELF-ANY). The remainder of an entry contains the\
\ sequence as lines following the id line up until a line starting\
\ with '>' or the end of the file. Whitespace and newlines are ignored.\
\ Characters 'A', 'T', 'G', 'C', 'a', 't', 'g', 'c' are retained\
\ and any other character is converted to 'N' (with the consequence\
\ that any IUB / IUPAC codes for ambiguous bases are converted to\
\ 'N'). There are no restrictions on line length. \n An empty value\
\ for this parameter indicates that no library should be used.",
prompt: Primer3 internal oligo mishybridizing library file (optional),
format: '("", " -mishyblibraryfile=" + str(value))[value is not None]',
argpos: 5, name: mishyblibraryfile, precond: hybridprobe, command: false}
- !mobyle/inputprogramparameter {comment: "The name of a file containing a\
\ nucleotide sequence library of sequences to avoid amplifying (for\
\ example repetitive sequences, or possibly the sequences of genes\
\ in a gene family that should not be amplified.) The file must be\
\ in (a slightly restricted) FASTA format (W. B. Pearson and D.J.\
\ Lipman, PNAS 85:8 pp 2444-2448 [1988]); we briefly discuss the\
\ organization of this file below. If this parameter is specified\
\ then EPrimer3 locally aligns each candidate primer against each\
\ library sequence and rejects those primers for which the local\
\ alignment score times a specified weight (see below) exceeds MAX-MISPRIMING.\
\ (The maximum value of the weight is arbitrarily set to 100.0.)\
\ \n Each sequence entry in the FASTA-format file must begin with\
\ an 'id line' that starts with '>'. The contents of the id line\
\ is 'slightly restricted' in that EPrimer3 parses everything after\
\ any optional asterisk ('*') as a floating point number to use as\
\ the weight mentioned above. If the id line contains no asterisk\
\ then the weight defaults to 1.0. The alignment scoring system used\
\ is the same as for calculating complementarity among oligos (e.g.\
\ SELF-ANY). The remainder of an entry contains the sequence as \
\ lines following the id line up until a line starting with '>' or\
\ the end of the file. Whitespace and newlines are ignored. Characters\
\ 'A', 'T', 'G', 'C', 'a', 't', 'g', 'c' are retained and any other\
\ character is converted to 'N' (with the consequence that any IUB\
\ / IUPAC codes for ambiguous bases are converted to 'N'). There\
\ are no restrictions on line length. \n An empty value for this\
\ parameter indicates that no repeat library should be used.", prompt: Primer3
mispriming library file (optional), format: '("", " -mispriminglibraryfile="
+ str(value))[value is not None]', argpos: 6, name: mispriminglibraryfile,
command: false}
- !mobyle/inputprogramparagraph
prompt: Additional section
name: additional
children:
- !mobyle/inputprogramparagraph
prompt: Program options
name: programsection
children:
- !mobyle/inputprogramparameter
comment: 'The maximum number of primer pairs to return. Primer pairs
returned are sorted by their ''quality'', in other words by the
value of the objective function (where a lower number indicates
a better primer pair). Caution: setting this parameter to a large
value will increase running time.'
prompt: Number of results to return (value greater than or equal to
0)
format: ("", " -numreturn=" + str(value))[value is not None and value!=vdef]
argpos: 7
name: numreturn
command: false
type: !mobyle/integertype {default: 5}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparagraph
prompt: Sequence options
name: seqoptsection
children:
- !mobyle/inputprogramparameter
comment: "A sub-region of the given sequence in which to pick primers.\
\ For example, often the first dozen or so bases of a sequence\
\ are vector, and should be excluded from consideration. The\
\ value for this parameter has the form \n (start),(end) \n \
\ where (start) is the index of the first base to consider, and\
\ (end) is the last in the primer-picking region."
prompt: Included region(s)
format: ("", " -includedregion=" + str(value))[value is not None]
argpos: 8
name: includedregion
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparameter
comment: "If one or more Targets is specified then a legal primer\
\ pair must flank at least one of them. A Target might be a simple\
\ sequence repeat site (for example a CA repeat) or a single-base-pair\
\ polymorphism. The value should be a space-separated list of\
\ \n (start),(end) \n pairs where (start) is the index of the\
\ first base of a Target, and (end) is the last \n E.g. 50,51\
\ requires primers to surround the 2 bases at positions 50 and\
\ 51."
prompt: Target region(s)
format: ("", " -targetregion=" + str(value))[value is not None]
argpos: 9
name: targetregion
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparameter
comment: "Primer oligos may not overlap any region specified in this\
\ tag. The associated value must be a space-separated list of\
\ \n (start),(end) \n pairs where (start) is the index of the\
\ first base of the excluded region, and and (end) is the last.\
\ This tag is useful for tasks such as excluding regions of low\
\ sequence quality or for excluding regions containing repetitive\
\ elements such as ALUs or LINEs. \n E.g. 401,407 68,70 forbids\
\ selection of primers in the 7 bases starting at 401 and the\
\ 3 bases at 68."
prompt: Excluded region(s)
format: ("", " -excludedregion=" + str(value))[value is not None]
argpos: 10
name: excludedregion
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparameter
comment: The sequence of a forward primer to check and around which
to design reverse primers and optional internal oligos. Must
be a substring of SEQUENCE.
prompt: Forward input primer sequence to check
format: ("", " -forwardinput=" + str(value))[value is not None]
argpos: 11
name: forwardinput
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparameter
comment: The sequence of a reverse primer to check and around which
to design forward primers and optional internal oligos. Must
be a substring of the reverse strand of SEQUENCE.
prompt: Reverse input primer sequence to check
format: ("", " -reverseinput=" + str(value))[value is not None]
argpos: 12
name: reverseinput
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparagraph
prompt: Primer options
name: primersection
children:
- !mobyle/inputprogramparameter
comment: Require the specified number of consecutive Gs and Cs at
the 3' end of both the forward and reverse primer. (This parameter
has no effect on the internal oligo if one is requested.)
prompt: Gc clamp (value greater than or equal to 0)
format: ("", " -gcclamp=" + str(value))[value is not None and value!=vdef]
argpos: 13
name: gcclamp
precond: primer
command: false
type: !mobyle/integertype {default: 0}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: Optimum length (in bases) of a primer oligo. EPrimer3 will
attempt to pick primers close to this length.
prompt: Primer optimum size (value greater than or equal to 0)
format: ("", " -osize=" + str(value))[value is not None and value!=vdef]
argpos: 14
name: osize
precond: primer
command: false
type: !mobyle/integertype {default: 20}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: Minimum acceptable length of a primer. Must be greater than
0 and less than or equal to MAX-SIZE.
prompt: Primer minimum size (value greater than or equal to 1)
format: ("", " -minsize=" + str(value))[value is not None and value!=vdef]
argpos: 15
name: minsize
precond: primer
command: false
type: !mobyle/integertype {default: 18}
ctrls:
- message: Value greater than or equal to 1 is required
test:
value: {'#gte': '1'}
- !mobyle/inputprogramparameter
comment: Maximum acceptable length (in bases) of a primer. Currently
this parameter cannot be larger than 35. This limit is governed
by the maximum oligo size for which EPrimer3's melting-temperature
is valid.
prompt: Primer maximum size (value less than or equal to 35)
format: ("", " -maxsize=" + str(value))[value is not None and value!=vdef]
argpos: 16
name: maxsize
precond: primer
command: false
type: !mobyle/integertype {default: 27}
ctrls:
- message: Value less than or equal to 35 is required
test:
value: {'#lte': '35'}
- !mobyle/inputprogramparameter
comment: Optimum melting temperature(Celsius) for a primer oligo.
EPrimer3 will try to pick primers with melting temperatures are close
to this temperature. The oligo melting temperature formula in
EPrimer3 is that given in Rychlik, Spencer and Rhoads, Nucleic
Acids Research, vol 18, num 21, pp 6409-6412 and Breslauer, Frank,
Bloecker and Marky, Proc. Natl. Acad. Sci. USA, vol 83, pp 3746-3750.
Please refer to the former paper for background discussion.
prompt: Primer optimum tm
format: ("", " -otm=" + str(value))[value is not None and value!=vdef]
argpos: 17
name: otm
precond: primer
command: false
type: !mobyle/floattype {default: 60.0}
- !mobyle/inputprogramparameter
comment: Minimum acceptable melting temperature(Celsius) for a primer
oligo.
prompt: Primer minimum tm
format: ("", " -mintm=" + str(value))[value is not None and value!=vdef]
argpos: 18
name: mintm
precond: primer
command: false
type: !mobyle/floattype {default: 57.0}
- !mobyle/inputprogramparameter
comment: Maximum acceptable melting temperature(Celsius) for a primer
oligo.
prompt: Primer maximum tm
format: ("", " -maxtm=" + str(value))[value is not None and value!=vdef]
argpos: 19
name: maxtm
precond: primer
command: false
type: !mobyle/floattype {default: 63.0}
- !mobyle/inputprogramparameter
comment: Maximum acceptable (unsigned) difference between the melting
temperatures of the forward and reverse primers.
prompt: Maximum difference in tm of primers
format: ("", " -maxdifftm=" + str(value))[value is not None and value!=vdef]
argpos: 20
name: maxdifftm
precond: primer
command: false
type: !mobyle/floattype {default: 100.0}
- !mobyle/inputprogramparameter
comment: Primer optimum GC percent.
prompt: Primer optimum gc percent
format: ("", " -ogcpercent=" + str(value))[value is not None and value!=vdef]
argpos: 21
name: ogcpercent
precond: primer
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: Minimum allowable percentage of Gs and Cs in any primer.
prompt: Primer minimum gc percent
format: ("", " -mingc=" + str(value))[value is not None and value!=vdef]
argpos: 22
name: mingc
precond: primer
command: false
type: !mobyle/floattype {default: 20.0}
- !mobyle/inputprogramparameter
comment: Maximum allowable percentage of Gs and Cs in any primer generated
by Primer.
prompt: Primer maximum gc percent
format: ("", " -maxgc=" + str(value))[value is not None and value!=vdef]
argpos: 23
name: maxgc
precond: primer
command: false
type: !mobyle/floattype {default: 80.0}
- !mobyle/inputprogramparameter
comment: The millimolar concentration of salt (usually KCl) in the
PCR. EPrimer3 uses this argument to calculate oligo melting temperatures.
prompt: Salt concentration (mm)
format: ("", " -saltconc=" + str(value))[value is not None and value!=vdef]
argpos: 24
name: saltconc
precond: primer
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: "The nanomolar concentration of annealing oligos in the PCR.\
\ EPrimer3 uses this argument to calculate oligo melting temperatures.\
\ The default (50nM) works well with the standard protocol used\
\ at the Whitehead/MIT Center for Genome Research--0.5 microliters\
\ of 20 micromolar concentration for each primer oligo in a 20\
\ microliter reaction with 10 nanograms template, 0.025 units/microliter\
\ Taq polymerase in 0.1 mM each dNTP, 1.5mM MgCl2, 50mM KCl,\
\ 10mM Tris-HCL (pH 9.3) using 35 cycles with an annealing temperature\
\ of 56 degrees Celsius. This parameter corresponds to 'c' in\
\ Rychlik, Spencer and Rhoads' equation (ii) (Nucleic Acids Research,\
\ vol 18, num 21) where a suitable value (for a lower initial\
\ concentration of template) is 'empirically determined'. The\
\ value of this parameter is less than the actual concentration\
\ of oligos in the reaction because it is the concentration of\
\ annealing oligos, which in turn depends on the amount of template\
\ (including PCR product) in a given cycle. This concentration\
\ increases a great deal during a PCR; fortunately PCR seems\
\ quite robust for a variety of oligo melting temperatures. \n\
\ See ADVICE FOR PICKING PRIMERS."
prompt: Dna concentration (nm)
format: ("", " -dnaconc=" + str(value))[value is not None and value!=vdef]
argpos: 25
name: dnaconc
precond: primer
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: The maximum allowable length of a mononucleotide repeat in
a primer, for example AAAAAA.
prompt: Maximum polynucleotide repeat (value greater than or equal
to 0)
format: ("", " -maxpolyx=" + str(value))[value is not None and value!=vdef]
argpos: 26
name: maxpolyx
precond: primer
command: false
type: !mobyle/integertype {default: 5}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparagraph
prompt: Product options
name: productsection
children:
- !mobyle/inputprogramparameter
comment: The optimum size for the PCR product. 0 indicates that there
is no optimum product size.
prompt: Product optimum size (value greater than or equal to 0)
format: ("", " -psizeopt=" + str(value))[value is not None and value!=vdef]
argpos: 27
name: psizeopt
precond: primer
command: false
type: !mobyle/integertype {default: 200}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: "The associated values specify the lengths of the product\
\ that the user wants the primers to create, and is a space separated\
\ list of elements of the form \n (x)-(y) \n where an (x)-(y)\
\ pair is a legal range of lengths for the product. For example,\
\ if one wants PCR products to be between 100 to 150 bases (inclusive)\
\ then one would set this parameter to 100-150. If one desires\
\ PCR products in either the range from 100 to 150 bases or in\
\ the range from 200 to 250 bases then one would set this parameter\
\ to 100-150 200-250. \n EPrimer3 favors ranges to the left side\
\ of the parameter string. EPrimer3 will return legal primers\
\ pairs in the first range regardless the value of the objective\
\ function for these pairs. Only if there are an insufficient\
\ number of primers in the first range will EPrimer3 return primers\
\ in a subsequent range."
prompt: Product size range
format: ("", " -prange=" + str(value))[value is not None and value!=vdef]
argpos: 28
name: prange
precond: primer
command: false
type: !mobyle/stringtype {default: 100-300}
- !mobyle/inputprogramparameter
comment: The optimum melting temperature for the PCR product. 0 indicates
that there is no optimum temperature.
prompt: Product optimum tm
format: ("", " -ptmopt=" + str(value))[value is not None and value!=vdef]
argpos: 29
name: ptmopt
precond: primer
command: false
type: !mobyle/floattype {default: 0.0}
- !mobyle/inputprogramparameter
comment: The minimum allowed melting temperature of the amplicon.
Please see the documentation on the maximum melting temperature of
the product for details.
prompt: Product minimum tm
format: ("", " -ptmmin=" + str(value))[value is not None and value!=vdef]
argpos: 30
name: ptmmin
precond: primer
command: false
type: !mobyle/floattype {default: -1000000.0}
- !mobyle/inputprogramparameter
comment: "The maximum allowed melting temperature of the amplicon.\
\ Product Tm is calculated using the formula from Bolton and \
\ McCarthy, PNAS 84:1390 (1962) as presented in Sambrook, Fritsch\
\ and Maniatis, Molecular Cloning, p 11.46 (1989, CSHL Press).\
\ \n Tm = 81.5 + 16.6(log10([Na+])) + .41*(%GC) - 600/length\
\ \n Where [Na+} is the molar sodium concentration, (%GC) is\
\ the percent of Gs and Cs in the sequence, and length is the\
\ length of the sequence. \n A similar formula is used by the\
\ prime primer selection program in GCG, which instead uses 675.0/length\
\ in the last term (after F. Baldino, Jr, M.-F. Chesselet, and\
\ M.E. Lewis, Methods in Enzymology 168:766 (1989) eqn (1) on\
\ page 766 without the mismatch and formamide terms). The formulas\
\ here and in Baldino et al. assume Na+ rather than K+. According\
\ to J.G. Wetmur, Critical Reviews in BioChem. and Mol. Bio.\
\ 26:227 (1991) 50 mM K+ should be equivalent in these formulae\
\ to .2 M Na+. EPrimer3 uses the same salt concentration value\
\ for calculating both the primer melting temperature and the\
\ oligo melting temperature. If you are planning to use the PCR\
\ product for hybridization later this behavior will not give\
\ you the Tm under hybridization conditions."
prompt: Product maximum tm
format: ("", " -ptmmax=" + str(value))[value is not None and value!=vdef]
argpos: 31
name: ptmmax
precond: primer
command: false
type: !mobyle/floattype {default: 1000000.0}
- !mobyle/inputprogramparagraph
prompt: Internal oligo input
name: oligosinput
children:
- !mobyle/inputprogramparameter
comment: "Middle oligos may not overlap any region specified by this\
\ tag. The associated value must be a space-separated list of\
\ \n (start),(end) \n pairs, where (start) is the index of\
\ the first base of an excluded region, and (end) is the last.\
\ Often one would make Target regions excluded regions for internal\
\ oligos."
prompt: Internal oligo excluded region
format: ("", " -oexcludedregion=" + str(value))[value is not None]
argpos: 32
name: oexcludedregion
precond: hybridprobe
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparameter
comment: The sequence of an internal oligo to check and around which
to design forward and reverse primers. Must be a substring of
SEQUENCE.
prompt: Internal oligo input sequence (if any)
format: ("", " -oligoinput=" + str(value))[value is not None]
argpos: 33
name: oligoinput
precond: hybridprobe
command: false
type: !mobyle/stringtype {}
- !mobyle/inputprogramparagraph
prompt: Internal oligo options
name: oligosection
children:
- !mobyle/inputprogramparameter
comment: Optimum length (in bases) of an internal oligo. EPrimer3
will attempt to pick primers close to this length.
prompt: Internal oligo optimum size (value greater than or equal to
0)
format: ("", " -osizeopt=" + str(value))[value is not None and value!=vdef]
argpos: 34
name: osizeopt
precond: hybridprobe
command: false
type: !mobyle/integertype {default: 20}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: Minimum acceptable length of an internal oligo. Must be greater
than 0 and less than or equal to INTERNAL-OLIGO-MAX-SIZE.
prompt: Internal oligo minimum size (value greater than or equal to
0)
format: ("", " -ominsize=" + str(value))[value is not None and value!=vdef]
argpos: 35
name: ominsize
precond: hybridprobe
command: false
type: !mobyle/integertype {default: 18}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: Maximum acceptable length (in bases) of an internal oligo.
Currently this parameter cannot be larger than 35. This limit
is governed by maximum oligo size for which EPrimer3's melting-temperature
is valid.
prompt: Internal oligo maximum size (value less than or equal to 35)
format: ("", " -omaxsize=" + str(value))[value is not None and value!=vdef]
argpos: 36
name: omaxsize
precond: hybridprobe
command: false
type: !mobyle/integertype {default: 27}
ctrls:
- message: Value less than or equal to 35 is required
test:
value: {'#lte': '35'}
- !mobyle/inputprogramparameter
comment: Optimum melting temperature (Celsius) for an internal oligo.
EPrimer3 will try to pick oligos with melting temperatures that
are close to this temperature. The oligo melting temperature
formula in EPrimer3 is that given in Rychlik, Spencer and Rhoads,
Nucleic Acids Research, vol 18, num 21, pp 6409-6412 and Breslauer,
Frank, Bloecker and Marky, Proc. Natl. Acad. Sci. USA, vol 83,
pp 3746-3750. Please refer to the former paper for background
discussion.
prompt: Internal oligo optimum tm
format: ("", " -otmopt=" + str(value))[value is not None and value!=vdef]
argpos: 37
name: otmopt
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 60.0}
- !mobyle/inputprogramparameter
comment: Minimum acceptable melting temperature(Celsius) for an internal
oligo.
prompt: Internal oligo minimum tm
format: ("", " -otmmin=" + str(value))[value is not None and value!=vdef]
argpos: 38
name: otmmin
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 57.0}
- !mobyle/inputprogramparameter
comment: Maximum acceptable melting temperature (Celsius) for an internal
oligo.
prompt: Internal oligo maximum tm
format: ("", " -otmmax=" + str(value))[value is not None and value!=vdef]
argpos: 39
name: otmmax
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 63.0}
- !mobyle/inputprogramparameter
comment: Internal oligo optimum GC percent.
prompt: Internal oligo optimum gc percent
format: ("", " -ogcopt=" + str(value))[value is not None and value!=vdef]
argpos: 40
name: ogcopt
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: Minimum allowable percentage of Gs and Cs in an internal
oligo.
prompt: Internal oligo minimum gc
format: ("", " -ogcmin=" + str(value))[value is not None and value!=vdef]
argpos: 41
name: ogcmin
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 20.0}
- !mobyle/inputprogramparameter
comment: Maximum allowable percentage of Gs and Cs in any internal
oligo generated by Primer.
prompt: Internal oligo maximum gc
format: ("", " -ogcmax=" + str(value))[value is not None and value!=vdef]
argpos: 42
name: ogcmax
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 80.0}
- !mobyle/inputprogramparameter
comment: The millimolar concentration of salt (usually KCl) in the
hybridization. EPrimer3 uses this argument to calculate internal
oligo melting temperatures.
prompt: Internal oligo salt concentration (mm)
format: ("", " -osaltconc=" + str(value))[value is not None and value!=vdef]
argpos: 43
name: osaltconc
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: The nanomolar concentration of annealing internal oligo in
the hybridization.
prompt: Internal oligo dna concentration (nm)
format: ("", " -odnaconc=" + str(value))[value is not None and value!=vdef]
argpos: 44
name: odnaconc
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 50.0}
- !mobyle/inputprogramparameter
comment: "The maximum allowable local alignment score when testing\
\ an internal oligo for (local) self-complementarity. Local self-complementarity\
\ is taken to predict the tendency of oligos to anneal to themselves\
\ The scoring system gives 1.00 for complementary bases, -0.25\
\ for a match of any base (or N) with an N, -1.00 for a mismatch,\
\ and -2.00 for a gap. Only single-base-pair gaps are allowed.\
\ For example, the alignment \n 5' ATCGNA 3' \n || | | \n 3'\
\ TA-CGT 5' \n is allowed (and yields a score of 1.75), but the\
\ alignment \n 5' ATCCGNA 3' \n || | | \n 3' TA--CGT 5' \n\
\ is not considered. Scores are non-negative, and a score of\
\ 0.00 indicates that there is no reasonable local alignment\
\ between two oligos."
prompt: Internal oligo maximum self complementarity (value less than
or equal to 9999.99)
format: ("", " -oanyself=" + str(value))[value is not None and value!=vdef]
argpos: 45
name: oanyself
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 12.0}
ctrls:
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparameter
comment: "The maximum allowable 3'-anchored global alignment score\
\ when testing a single oligo for self-complementarity. \n The\
\ scoring system is as for the Maximum Complementarity argument.\
\ In the examples above the scores are 7.00 and 6.00 respectively.\
\ Scores are non-negative, and a score of 0.00 indicates that\
\ there is no reasonable 3'-anchored global alignment between\
\ two oligos. In order to estimate 3'-anchored global alignments\
\ for candidate oligos, Primer assumes that the sequence from\
\ which to choose oligos is presented 5' to 3'. \n INTERNAL-OLIGO-SELF-END\
\ is meaningless when applied to internal oligos used for hybridization-based\
\ detection, since primer-dimer will not occur. We recommend\
\ that INTERNAL-OLIGO-SELF-END be set at least as high as INTERNAL-OLIGO-SELF-ANY."
prompt: Internal oligo maximum 3' self complementarity (value less
than or equal to 9999.99)
format: ("", " -oendself=" + str(value))[value is not None and value!=vdef]
argpos: 46
name: oendself
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 12.0}
ctrls:
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparameter
comment: The maximum allowable length of an internal oligo mononucleotide
repeat, for example AAAAAA.
prompt: Internal oligo maximum polynucleotide repeat (value greater
than or equal to 0)
format: ("", " -opolyxmax=" + str(value))[value is not None and value!=vdef]
argpos: 47
name: opolyxmax
precond: hybridprobe
command: false
type: !mobyle/integertype {default: 5}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: Similar to MAX-MISPRIMING except that this parameter applies
to the similarity of candidate internal oligos to the library
specified in INTERNAL-OLIGO-MISHYB-LIBRARY.
prompt: Internal oligo maximum mishybridization (value less than or
equal to 9999.99)
format: ("", " -omishybmax=" + str(value))[value is not None and value!=vdef]
argpos: 48
name: omishybmax
precond: hybridprobe
command: false
type: !mobyle/floattype {default: 12.0}
ctrls:
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparagraph
prompt: Advanced section
name: advanced
children:
- !mobyle/inputprogramparameter
comment: If this flag is true, produce LEFT-EXPLAIN, RIGHT-EXPLAIN, and
INTERNAL-OLIGO-EXPLAIN output tags, which are intended to provide
information on the number of oligos and primer pairs that EPrimer3
examined, and statistics on the number discarded for various reasons.
prompt: Explain flag
format: ("", " -explainflag")[ bool(value) ]
argpos: 49
name: explainflag
command: false
type: !mobyle/booleantype {default: false}
- !mobyle/inputprogramparameter
comment: If the associated value is true, then EPrimer3 creates two output
files for each input SEQUENCE. File (sequence-id).for lists all acceptable
forward primers for (sequence-id), and (sequence-id).rev lists all
acceptable reverse primers for (sequence-id), where (sequence-id)
is the value of the SEQUENCE-ID tag (which must be supplied). In
addition, if the input tag TASK is 1 or 4, EPrimer3 produces a file
(sequence-id).int, which lists all acceptable internal oligos.
prompt: Create results files for each sequence
format: ("", " -fileflag")[ bool(value) ]
argpos: 50
name: fileflag
command: false
type: !mobyle/booleantype {default: false}
- !mobyle/inputprogramparameter
comment: This parameter is the index of the first base in the input sequence.
For input and output using 1-based indexing (such as that used in
GenBank and to which many users are accustomed) set this parameter
to 1. For input and output using 0-based indexing set this parameter
to 0. (This parameter also affects the indexes in the contents of
the files produced when the primer file flag is set.)
prompt: First base index
format: ("", " -firstbaseindex=" + str(value))[value is not None and value!=vdef]
argpos: 51
name: firstbaseindex
command: false
type: !mobyle/integertype {default: 1}
- !mobyle/inputprogramparameter
comment: If true pick a primer pair even if LEFT-INPUT, RIGHT-INPUT, or
INTERNAL-OLIGO-INPUT violates specific constraints.
prompt: Pick anyway
format: ("", " -pickanyway")[ bool(value) ]
argpos: 52
name: pickanyway
command: false
type: !mobyle/booleantype {default: false}
- !mobyle/inputprogramparameter
comment: The maximum allowed weighted similarity with any sequence in
MISPRIMING-LIBRARY.
prompt: Primer maximum mispriming (value less than or equal to 9999.99)
format: ("", " -maxmispriming=" + str(value))[value is not None and value!=vdef]
argpos: 53
name: maxmispriming
command: false
type: !mobyle/floattype {default: 12.0}
ctrls:
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparameter
comment: The maximum allowed sum of weighted similarities of a primer
pair (one similarity for each primer) with any single sequence in
MISPRIMING-LIBRARY.
prompt: Primer pair maximum mispriming (value less than or equal to 9999.99)
format: ("", " -pairmaxmispriming=" + str(value))[value is not None and
value!=vdef]
argpos: 54
name: pairmaxmispriming
command: false
type: !mobyle/floattype {default: 24.0}
ctrls:
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparameter
comment: Maximum number of unknown bases (N) allowable in any primer.
prompt: Maximum ns accepted in a primer (value greater than or equal to
0)
format: ("", " -numnsaccepted=" + str(value))[value is not None and value!=vdef]
argpos: 55
name: numnsaccepted
command: false
type: !mobyle/integertype {default: 0}
ctrls:
- message: Value greater than or equal to 0 is required
test:
value: {'#gte': '0'}
- !mobyle/inputprogramparameter
comment: "The maximum allowable local alignment score when testing a single\
\ primer for (local) self-complementarity and the maximum allowable\
\ local alignment score when testing for complementarity between\
\ forward and reverse primers. Local self-complementarity is taken\
\ to predict the tendency of primers to anneal to each other without\
\ necessarily causing self-priming in the PCR. The scoring system\
\ gives 1.00 for complementary bases, -0.25 for a match of any base\
\ (or N) with an N, -1.00 for a mismatch, and -2.00 for a gap. Only\
\ single-base-pair gaps are allowed. For example, the alignment \n\
\ 5' ATCGNA 3' \n ...|| | | \n 3' TA-CGT 5' \n is allowed (and\
\ yields a score of 1.75), but the alignment \n 5' ATCCGNA 3' \n\
\ ...|| | | \n 3' TA--CGT 5' \n is not considered. Scores are non-negative,\
\ and a score of 0.00 indicates that there is no reasonable local\
\ alignment between two oligos."
prompt: Maximum self complementarity (value from 0.00 to 9999.99)
format: ("", " -selfany=" + str(value))[value is not None and value!=vdef]
argpos: 56
name: selfany
command: false
type: !mobyle/floattype {default: 8.0}
ctrls:
- message: Value greater than or equal to 0.00 is required
test:
value: {'#gte': '0.0'}
- message: Value less than or equal to 9999.99 is required
test:
value: {'#lte': '9999.99'}
- !mobyle/inputprogramparameter
comment: "The maximum allowable 3'-anchored global alignment score when\
\ testing a single primer for self-complementarity, and the maximum\
\ allowable 3'-anchored global alignment score when testing for complementarity\
\ between forward and reverse primers. The 3'-anchored global alignment\
\ score is taken to predict the likelihood of PCR-priming primer-dimers,\
\ for example \n 5' ATGCCCTAGCTTCCGGATG 3' \n .............||| |||||\
\ \n ..........3' AAGTCCTACATTTAGCCTAGT 5' \n or \n 5' AGGCTATGGGCCTCGCGA\
\ 3' \n ...............|||||| \n ............3' AGCGCTCCGGGTATCGGA\
\ 5' \n The scoring system is as for the Maximum Complementarity\
\ argument. In the examples above the scores are 7.00 and 6.00 respectively.\
\ Scores are non-negative, and a score of 0.00 indicates that there\
\ is no reasonable 3'-anchored global alignment between two oligos.\
\ In order to estimate 3'-anchored global alignments for candidate\
\ primers and primer pairs, Primer assumes that the sequence from\
\ which to choose primers is presented 5' to 3'. It is nonsensical\
\ to provide a larger value for this parameter than for the Maximum\
\ (local) Complementarity parameter because the score of a local\
\ alignment will always be at least as great as the score of a global\
\ alignment."
prompt: Maximum 3' self complementarity (value greater than or equal to
0.00)
format: ("", " -selfend=" + str(value))[value is not None and value!=vdef]
argpos: 57
name: selfend
command: false
type: !mobyle/floattype {default: 3.0}
ctrls:
- message: Value greater than or equal to 0.00 is required
test:
value: {'#gte': '0.0'}
- !mobyle/inputprogramparagraph
prompt: Primer penalty weights
name: primerweights
children:
- !mobyle/inputprogramparameter
comment: 'The maximum stability for the five 3'' bases of a forward
or reverse primer. Bigger numbers mean more stable 3'' ends. The value
is the maximum delta G for duplex disruption for the five 3''
bases as calculated using the nearest neighbor parameters published
in Breslauer, Frank, Bloecker and Marky, Proc. Natl. Acad. Sci.
USA, vol 83, pp 3746-3750. EPrimer3 uses a completely permissive
default value for backward compatibility (which we may change
in the next release). Rychlik recommends a maximum value of 9
(Wojciech Rychlik, ''Selection of Primers for Polymerase Chain
Reaction'' in BA White, Ed., ''Methods in Molecular Biology,
Vol. 15: PCR Protocols: Current Methods and Applications'', 1993,
pp 31-40, Humana Press, Totowa NJ).'
prompt: Maximum 3' end stability (value less than or equal to 999.9999)
format: ("", " -maxendstability=" + str(value))[value is not None
and value!=vdef]
argpos: 58
name: maxendstability
command: false
type: !mobyle/floattype {default: 9.0}
ctrls:
- message: Value less than or equal to 999.9999 is required
test:
value: {'#lte': '999.9999'}
- !mobyle/inputprogramparagraph
prompt: Output section
name: output
children:
- !mobyle/inputprogramparameter
prompt: Name of the output file (outfile)
format: ("" , " -outfile=" + str(value))[value is not None]
argpos: 59
name: outfile
command: false
type: !mobyle/stringtype {default: eprimer3.outfile}
- !mobyle/inputprogramparameter
prompt: Turn off any prompting
format: '" -auto -stdout"'
argpos: 60
name: auto
command: false
hidden: true
type: !mobyle/stringtype {}
outputs: !mobyle/outputparagraph
children:
- !mobyle/outputprogramparagraph
prompt: Output section
name: output
children:
- !mobyle/outputprogramparameter
prompt: Outfile_out option
filenames: outfile
name: outfile_out
type: !mobyle/formattedtype
data_terms: ['EDAM_data:2048']
- !mobyle/outputprogramparameter
prompt: Standard output
filenames: '"eprimer3.out"'
name: stdout
output_type: stdout
type: !mobyle/formattedtype
data_terms: ['EDAM_data:2048']
- !mobyle/outputprogramparameter
prompt: Standard error
filenames: '"eprimer3.err"'
name: stderr
type: !mobyle/formattedtype
data_terms: ['EDAM_data:2048']
operations: ['EDAM_operation:0308', 'EDAM_operation:0309']
topics: ['EDAM_topic:0195']
documentation_links: ['http://bioweb2.pasteur.fr/docs/EMBOSS/eprimer3.html', 'http://emboss.sourceforge.net/docs/themes']
command: eprimer3
env: {}