Modified oligonucleotides and applications thereof

A purified single-stranded polynucleotide primer comprising at least one modified pyrimidine nucleobase of the structure

• wherein
• X is N or C,
• R1 is selected from —H, —F, —Cl, —Br, C1-C6 alkyl, C1-C6 substituted alkyl, C3-C10 aryl, C3-C10 substituted aryl, —CF3, —CF2H, —CF2CH3, —CF2CF3, —CCl3, —CN, —CHO, —CO2R, —SO3R, —PO3RR, —C(O)NRR, azido, and —NO2 where each R is independently —H, C1-C6 alkyl or C3-C10 aryl or alkylaryl, and
• R2 is selected from —H, —F, —Cl, —Br, C1-C6 alkyl, C1-C6 substituted alkyl, C3-C10 aryl, C3-C10 substituted aryl, —CF3, —CF2H, —CF2CH3, —CF2CF3, —CCl3, —CN, —CHO, —CO2R, —SO3R, —PO3RR, —C(O)NRR, azido, and —NO2 where each R is independently H, C1-C6 alkyl or C3-C10 aryl or alkylaryl,
• wherein
• at least one of R1 or R2 is an electron withdrawing substituent or X is N, such that when X is N, R2 is absent,
• at least one said modified pyrimidine nucleobase is no more than 4 nucleotides from the 3′ terminus of the primer, and
• said purified single-stranded polynucleotide primer comprising said modified pyrimidine nucleobase now complementary to a target sequence.