to begin, I need synthesis single strand oligo or double strands oligo DNA?

I need synthesis single strand oligo or double complementary paired strands oligo DNA (designed for interesting target)?


Login or Signup to leave a comment
Ruby Wyrmabout 4 years ago

I usually order single strand oligos as when I am ordering primers, and resuspend them to 100uM.

Then I phosphorylate (using T4 PNK from NEB) and anneal them in the same reaction using the following protocol:

T4 PNK buffer 10x 1uL

top strand 100uM 1uL

bottom strand 100uM 1uL

T4 PNK enzyme 1uL

H20 6uL

Total 10uL

Incubate 30' at 37C (this is for phosphorylating the single strand oligos).

Then, 95C for 5 minutes (this will resolve any secondary structure the single strand oligos can make) and let the tube cool down to room temperature for at least 1 hour (this will allow the formation of some double stranded DNA).

The final concentration of your dsDNA will be considered 10uM and you will have to dilute it accordingly to fit to your ligation reaction (you have to dilute a lot).

Jade Nagaabout 4 years ago

thanks a lot

Find your community. Ask questions. Science is better when we troubleshoot together.
Find your community. Ask questions. Science is better when we troubleshoot together.

Have a question?

Contact support@scifind.net or check out our support page.