Photocleavable nucleotide reagents with high stability

A method of amplifying a target nucleic acid sequence in a sample comprising:

• providing amplification reagents to perform an amplifying step, wherein the amplification reagents comprise a modified nucleoside triphosphate having a photocleavable moiety and having a structure of:
• where Base is a purine or pyrimidine base or an analog;
• and
• wherein prior to the amplifying step, the modified nucleoside triphosphate is irradiated with light having a wavelength capable of cleaving the photocleavable moiety.

A method of reducing or preventing non-specific amplification of nucleic acid in a sample during amplification by polymerase chain reaction (PCR) comprising:

• a) providing amplification reagents that are used to amplify a target nucleic acid, wherein the amplification reagents comprise a nucleic acid polymerase and a modified nucleoside triphosphate having a photocleavable moiety and having a structure of:
• where Base is a purine or pyrimidine base or an analog;
• b) contacting the sample with the amplification reagents at room temperature, whereby the photocleavable moiety on the modified nucleoside triphosphate prevents processing by the nucleic acid polymerase, thereby suppressing formation of non-specific amplification products from room temperature activity of the nucleic acid polymerase;
• c) prior to amplifying, irradiating the modified nucleoside triphosphate with light having a wavelength capable of cleaving the photocleavable moiety to release free nucleoside triphosphate;
• and
• d) amplifying the target nucleic acid by PCR.