Analysis ----Primer3
Primer3   Pick primers from a DNA sequence.   
Paste source sequence below (5'->3', string of ACGTNacgtn -- other letters treated as N -- numbers and blanks ignored). FASTA format ok. Please N-out undesirable sequence (vector, ALUs, LINEs, etc.) or use a Mispriming Library (repeat library):
Pick left primer, or use left primer below:
Pick hybridization probe (internal oligo), or use oligo below:
Pick right primer, or use right primer below (5' to 3' on opposite strand):
Sequence Id: A string to identify your output.
Targets: E.g. 50,2 requires primers to surround the 2 bases at positions 50 and 51. Or mark the source sequence with [ and ]: e.g. ...ATCT[CCCC]TCAT.. means that primers must flank the central CCCC.
Excluded Regions: E.g. 401,7 68,3 forbids selection of primers in the 7 bases starting at 401 and the 3 bases at 68. Or mark the source sequence with < and >: e.g. ...ATCT<CCCC>TCAT.. forbids primers in the central CCCC.

General Primer Picking Conditions

Primer Size Min: Opt: Max:
Primer Tm Min: Opt: Max: Max Tm Difference:
Product Tm Min: Opt: Max:
Primer GC% Min: Opt: Max:
Salt Concentration: Annealing Oligo Concentration: (Not the concentration of oligos in the reaction mix but of those annealing to template.)
Liberal Base Show Debuging Info Do not treat ambiguity codes in libraries as consensus

Other Per-Sequence Inputs

Included Region: E.g. 20,400: only pick primers in the 400 base region starting at position 20. Or use { and } in the source sequence to mark the beginning and end of the included region: e.g. in ATC{TTC...TCT}AT the included region is TTC...TCT.

Objective Function Penalty Weights for Primers

Tm Lt: Gt:
Size Lt: Gt:
GC% Lt: Gt:

Objective Function Penalty Weights for Primer Pairs

Product Size Lt: Gt:
Product Tm Lt: Gt:

Hyb Oligo (Internal Oligo) Per-Sequence Inputs

Hyb Oligo (Internal Oligo) General Conditions

Hyb Oligo Size: Min Opt Max
Hyb Oligo Tm: Min Opt Max
Hyb Oligo GC% Min: Opt: Max:
Hyb Oligo Mishyb Library: Hyb Oligo Max Mishyb:

Objective Function Penalty Weights for Hyb Oligos (Internal Oligos)

Hyb Oligo Tm Lt: Gt:
Hyb Oligo Size Lt: Gt:
Hyb Oligo GC% Lt: Gt:
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