Remove FAM label from oligo, enzymatically or chemically ?

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Abhijeet Singh

I dont know how to remove the FAM label, but it would not be practical either. If a tag is cleaved from oligo, still it will be a mixture and would be hard to separate. It would be much cheaper to make new oligos without any tag.

Michiel Vanmeert

hi thank you for the answer.

The case is we have some modified (limited) in the oligo. The separation could be done by PAGE. The tag will just run off.

The cleavage would be the hardest part. Maybe by using AMA?

Thank you for all your help !

Paul Rutland

I suspect chemical removal may be difficult as the FAM has survived deprotection with hot concentrated ammonia in the manufacture process without any problems ( milder methods are used these days) and acid methods are likely to cause depurination but I am intrigued as to why you need to remove the label. I think many downstream processes will be unaffected by the label and even sequencing should be possible by analysing out the template FAM peak .Sorry not helpful to you just being nosey

The oligo is combined with another FAM labeled oligo. On a gel, I only want to see 1 of these 2 oligos. Would photobleaching be an option?

thanks!

I wondered about that but decided that UV irradiation might denature the oligo but it is worth a try. Naturally if you have a light source emitting at 494nm that should bleach without damaging the DNA so possibly leaving a sample on ice or lyophilised near a white light source is worth a try but as Abhijeet says oligos are pretty cheap these days

Thank you very much for your answer . The modified nucleotides in the oligo (more stable than DNA) make the oligo expensive, together with the limited amount to build them.

I will try with AMA at 60 degrees Celcius, and another sample irradiated.

Thanks !

ah pity there went all ideas of smaller oligos to amplify the 38mer without the dye ( asymmetric pcr). I would be interested to know how this one pans out

good luck

Paul