Using Amino-Modified Oligos
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A primary amino function can be incorporated during oligonucleotide
synthesis at either the 5'or 3' end or internally using an amino dT reagent.
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In order to successfully conjugate haptens or fluorophores to the primary amino
group, it is necessary to ensure that the oligonucleotide has sodium as the
counter-ion rather than ammonium, since the latter may interfere with the conjugation
reaction. Oligonucleotides purchased from MIDLAND will either be in the sodium or
ammonium form, depending on the type of purification. Gel filtration (GF) Grade
and Reverse Phase (RP) HPLC Grade oligonucleotides will be in the ammonium salt
form. Anion Exchange (AE) HPLC Grade and PAGE Grade oligonucleotides will be in
the sodium salt form.
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If an oligonucleotide is the ammonium salt form, it can be exchanged to
the sodium salt form quite simply. The lyophilized oligonucleotide should be
resuspended in a small volume of 0.2 M NaCl. The excess salt can then be removed
by gel filtration on Sephadex G25 or by ethanol precipitation of the oligonucleotide.
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