PrbbBB:Oligo Annealing

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Short to medium length (up to 120 bp) fragments of dsDNA can be generated directly from two complementary single-stranded oligonucleotides. Custom overhangs for ligation or In-Fusion can be introduced as well, simply by having some non-complementary ends. The two single-stranded Oligos then need to be annealed into a double-stranded fragment.

See also:

The idea is to heat up the mixture to close to 100 C and let it slowly cool down to room temperature over the course of one hour.


  • Thermocycler
  • some buffer


  1. step 1
  2. step 2


Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

raik: We are just starting to use this protocol -- share your experience!


Error fetching PMID 17293868:
  1. Error fetching PMID 17293868: [Li2007]


or instead, discuss this protocol.

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