Pydnaweb is a flask application that exposes some of the functionality of the pydna Python package as an online service.
It can be hosted on pythonanywhere for example.
Development of Pydna&Pydnaweb is led by Björn Johansson at the Department of Biology, University of Minho.
If you have question not answered below or suggestions, please ask in the Google group (preferred) or create an issue on GitHub.
All tools accept sequences in FASTA or Genbank format. The formats can be mixed.
WebPCR simulates PCR given at least two primers and a template sequence as a list.
The last of the sequences in this list is assumed to be the template sequence while all preceding sequences are assumed to be primers.
The example below, shows the template MyTemplate, ForwardPrimer and ReversePrimer.
>ForwardPrimer
gctactacacacgtactgactg
>ReversePrimer
tgtggttactgactctatcttg
>MyTemplate
gctactacacacgtactgactgcctccaagatagagtcagtaaccaca
A report is generated as detailed below:
Template MyTemplate 48 bp linear limit=16:
ForwardPrimer anneals forward (--->) at 22
ReversePrimer anneals reverse (<---) at 26
5gctactacacacgtactgactg...caagatagagtcagtaaccaca3
||||||||||||||||||||||
3gttctatctcagtcattggtgt5
5gctactacacacgtactgactg3
||||||||||||||||||||||
3cgatgatgtgtgcatgactgac...gttctatctcagtcattggtgt5
Taq DNA polymerase
|95°C|95°C | |tmf:62.4
|____|_____ 72°C|72°C|tmr:59.1
|3min|30s \ 48.7°C _____|____|45s/kb
| | \______/ 0:30|5min|GC 47%
| | 30s | |48bp
WebPCR can handle circular templates. A circular template can be indicated by the Genbank file on the LOCUS line or by supplying the keyword "circular" in the FASTA header like this:
>ForwardPrimer
gatagagtcagtaacc
>ReversePrimer
cagtcagtacgtgtgt
>MyTemplate circular
gctactacacacgtactgactgcctccaagatagagtcagtaaccaca
As the forward primer anneals after the reverse primer, no PCR product would be formed on a linear template. On a circular template, the amplification occurs across the origin of the sequence.
Template MyTemplate 48 bp circular limit=16:
ForwardPrimer anneals forward (--->) at 45
ReversePrimer anneals reverse (<---) at 6
5gatagagtcagtaacc...acacacgtactgactg3
||||||||||||||||
3tgtgtgcatgactgac5
5gatagagtcagtaacc3
||||||||||||||||
3ctatctcagtcattgg...tgtgtgcatgactgac5
Taq DNA polymerase
|95°C|95°C | |tmf:48.5
|____|_____ 72°C|72°C|tmr:53.7
|3min|30s \ 43.4°C _____|____|45s/kb
| | \______/ 0:30|5min|GC 46%
| | 30s | |41bp
Pfu-Sso7d DNA polymerase
|98°C|98°C | |tmf:42.1
|____|_____ 72°C|72°C|tmr:48.9
|30s |10s \ 45.1°C _____|____|15s/kb
| | \______/ 0:10|5min|GC 46%
| | 10s | |41bp
>41bp_PCR_prod
gatagagtcagtaaccacagctactacacacgtactgactg
Calculates the melting temperature for a list of primer sequences using data and a the Bio.SeqUtils.MeltingTemp.Tm_NN function.
There are many online primer melting calculators available online, but they rarely expose all the details of algorithms and data used.
For details on the decisions for the default values see here.
Designs primers for one or more template sequences. Three sequences are returned for each input sequence, representing the two primers and the original sequence, followed by a suggested PCR program.
>f40 12-mer
GTCCCCGAGGCG
>r40 19-mer
CTTCTACAAAACCGCGTCA
>seq
GTCCCCGAGGCGGACACTGATTGACGCGGTTTTGTAGAAG
Taq DNA polymerase
|95°C|95°C | |tmf:59.9
|____|_____ 72°C|72°C|tmr:59.8
|3min|30s \ 49.7°C _____|____|45s/kb
| | \______/ 0:30|5min|GC 57%
| | 30s | |40bp
>40bp_PCR_prod
GTCCCCGAGGCGGACACTGATTGACGCGGTTTTGTAGAAG
BamHI
EcoRI
SmaI
NotI
>ins1
GGATCCaaCCCGGGaGGATCC
>ins2
GGATCCttCCCGGGtGGATCC
>pUCmu 1669bp circular
ACGCGTCGCGAGGCCATATGGGTTAACCCATGGCCAAGCTTGCATGCCTGCAGGTCGACTCTAGAGGATCCCGGGTACCGAGCTCGAATTCGGATATCCTCGAGACTAGTGGGCCCGTTTAAACACATGTGTTTTTCCATAGGCTCCGCCCCCCTGACGAGCATCACAAAAATCGACGCTCAAGTCAGAGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGAAGCTCCCTCGTGCGCTCTCCTGTTCCGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGGGAAGCGTGGCGCTTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGTCGTTCGCTCCAAGCTGGGCTGTGTGCACGAACCCCCCGTTCAGCCCGACCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGTAAGACACGACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGATTAGCAGAGCGAGGTATGTAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACGGCTACACTAGAAGAACAGTATTTGGTATCTGCGCTCTGCTGAAGCCAGTTACCTTCGGAAAAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACCGCTGGTAGCGGTGGTTTTTTTGTTTGCAAGCAGCAGATTACGCGCAGAAAAAAAGGATCTCAAGAAGATCCTTTGATCTTTTCTACTACCAATGCTTAATCAGTGAGGCACCTATCTCAGCGATCTGTCTATTTCGTTCATCCATAGTTGCCTGACTCCCCGTCGTGTAGATAACTACGATACGGGAGGGCTTACCATCTGGCCCCAGTGCTGCAATGATACCGCGAGACCCACGCTCACCGGCTCCAGATTTATCAGCAATAAACCAGCCAGCCGGAAGGGCCGAGCGCAGAAGTGGTCCTGCAACTTTATCCGCCTCCATCCAGTCTATTAATTGTTGCCGGGAAGCTAGAGTAAGTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGCATCGTGGTGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTCCCAACGATCAAGGCGAGTTACATGATCCCCCATGTTGTGCAAAAAAGCGGTTAGCTCCTTCGGTCCTCCGATCGTTGTCAGAAGTAAGTTGGCCGCAGTGTTATCACTCATGGTTATGGCAGCACTGCATAATTCTCCTACTGTCATGCCATCCGTAAGATGCTTTTCTGTGACTGGTGAGTACTCAACCAAGTCATTCTGAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGCCCGGCGTCAATACGGGATAATACCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAAACGTTCTTCGGGGCGAAAACTCTCAAGGATCTTACCGCTGTTGAGATCCAGTTCGATGTAACCCACTCGTGCACCCAACTGATCTTCAGCATCTTTTACTTTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGCAAAATGCCGCAAAAAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCCTTTTTCAATATTATTGAAGCATTTATCAGGGTTATTGTCTCATGAGCGGATACATA