User GuidesSnapGene Designing SequencesAnneal Oligonucleotide Pairs

Anneal Oligonucleotide Pairs

How do I create a double strand sequence from two complementary primers?

Create a Sticky-Ended Double -Stranded Fragment from two Oligonucleotides

This technique simulates methods commonly used to generate short double-stranded linkers by annealing equimolar amounts of complementary oligonucleotides.

Click Actions → Anneal Oligos...

In this example the complementary primers have been designed to create a double stranded fragment with 5' overhangs compatible with BamHI and NotI restriction enzyme sites.

Enter the names and sequences of your primer pair into the fields provided.

SnapGene will automatically detect the overlap between the two primer sequences. Note that the overlapping regions must match perfectly for the overlap to be detected.

SnapGene will automatically detect overhangs and report any restriction enzymes that generate compatible cohesive ends. SnapGene will also report the melting temperature (Tm) calculated for the overlap between the complementary oligonucleotides.

Give the new "product" an appropriate name and click Anneal to create your double stranded sequence.

Show the Description Panel to add details about the new sequence.

Click File → Save to save the sequence to the appropriate location on your computer.

Anneal and Backfill Oligonucleotide Pairs

This technique simulates methods commonly used to synthesize short double-stranded linkers by annealing equimolar amounts of complementary oligonucleotides, and then backfilling with DNA polymerase.

Click Actions → Anneal Oligos...

In this example complementary primers create a double stranded fragment with 3' recessed ends suitable for backfilling with DNA polymerase

Enter the names and sequences of your primer pair into the fields provided.

SnapGene will automatically detect the overlap between the two primer sequences. Note the overlapping regions must match perfectly for the overlap to be detected.

SnapGene reports the melting temperature (Tm) for the overlap between the complementary oligonucleotides.

To simulate backfilling of 3' recessed ends by DNA polymerase, check the option to "Fill in 5' overhangs". The nucleotides added by simulated backfilling of overhangs are colored red.

Give the "product" an appropriate name, then click Anneal to create the new sequence.

Show the Description Panel to add details about the new sequence.

Click File → Save to save the new sequence to the appropriate location on your computer.