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- User primers - the pair of forward and reverse primers user can check on the absence of unwanted connections (like hairpins, self- and hetero-dimers).
- Choose generated sequences as user primer's end - user primers could be extended with the 8-bases sequences on 5' and 3' ends. These sequences do not form too simple unwanted connections (for example, "AAAATTTT", obviously, has a hairpin, so there is no such a sequence in the list).
- Parameters of priming sequences - the range of melting temperature, Gibbs free energy and length primers SHOULD have.
- Parameters to exclude in whole primers - the minimum melting temperature, the maximum Gibbs free energy and the maximum base pairs length which may be present in the self- and hetero-dimers in the considered primer.
- Select areas for priming search - select areas to search forward and reverse primers in.
- Open the backbone sequence - choose a sequence to be added to 5' or 3' end of the result primer.
- Other sequences in PCR reaction - the list of sequence to check the unwanted connections with. The unwanted connections should fit to "Parameters to exclude in whole primers".
Forward primer - forward primer.
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