I am doing DNA synthesis, and want to add EMCA in the 5' end via solid phase synthesis, but the DNA cleavage need to be done in ammonium hydroxide at 50 ℃. I want to ask will the amide bond between DNA and EMCA be broke during the cleavage?
Amide bonds do cleave in hydroxide solutions yes. You cant reflux DMF with KOH to get rid of water for that reason. However the rate of cleavage is dependent on [OH-] and temperature so I dont know to what extent the amide bond would break off the top of my head.
If you knew the [OH-] and rate of hydrolysis for a prototypical amide bond at room temperature, you could probably calculate the rate of hydrolysis at 50 °C
What is the structure of EMCA？ In ammonium hydroxide at 50 ℃ ， some amide bonds are stable， some are not！For example， Fam-NH-（CH2）6 - amide bond is stable。 F3CCONH-（CH2）6- amide bond is not stable。All NH2 protecting groups on A，G， C，are not stable。
The 5'-amino-DNA conjugated with COOH or COONHS, you do not need to use ammonium hydroxide at 50 ℃ anymore, because the 5'-amino-DNA should be already treated with ammonium hydroxide at 50 ℃ .
I guess that Amide bond is quite strong bond, and usually needs acidic conditions to be broken
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