Corum:DNA Hybridization

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Hybridization of complimentary ssDNA oligonucleotides to create dsDNA linker.


  • 5 μL 2mM sense oligonucleotide
  • 5 μL 2mM antisense oligonucleotide

NOTE: If X = nmol of lyophilized oligonucleotide, add X/2 μL water to obtain 2mM final concentration.


  1. Combine 5 μL sense oligonucleotide and 5 μL antisense oligonucleotide. Vortex and spin.
  2. Double seal tube airtight with parafilm and press firmly in weighted holder.
  3. Boil 800 mL water in a 1L beaker.
  4. Wait 1 min.
  5. Place weighted holder with sealed oligonucleotides into the beaker.
  6. Incubate on bench overnight or until water is about room temperature (~4 hr). At the end of this process, the complimentary ssDNA's are hybridized into as single dsDNA linker with sticky ends at 1mM final concentration.
  7. Perform two 1/100 dilutions (label all tubes clearly):
    • 2 μL 1mM linker + 198 μL H2O = 200 μL 10μM linker
    • 2 μL 10μM linker + 198 μL H2O = 200 μL 100nM linker
  8. Store 1mM, 10μM, and 100nM stocks at -40 °C in latest "Primer" box. Keep an aliquot of the 100nM linker in your own box for use in constructions.


  • SC 19:27, 24 July 2012 (EDT):


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