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Backbone sequence is the sequence that is added to 5' end of the primer for the following assembly of the PCR product with the backbone molecule. This parameter is optional. You may set the backbone sequence, the sequence end you want to add backbone to and the length of the sequence from 5' and 3' ends to check if these sequences have unwanted hairpins, self- and hetero-dimers. On the pictures below you may see the differences between "5' insert to 5' backbone" and "5' insert to 3' backbone" ("insert" in this case is the same as "primer").

5' insert to 5' backbone.Image Added

5' insert to 3' backbone.Image Added

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Type two primers for running In Silico PCR. If the primers pair is invalid for running the PCR process then the warning is shown. Also, primers for the running In silico PCR can be chosen from a primer library. Click the following button to choose a primer from the primers library: 

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