RNase H-based assays utilizing modified RNA monomers
View Patent ↗The present invention pertains to novel oligonucleotide compounds for use in various biological assays, such as nucleic acid amplification, ligation and sequencing reactions. The novel oligonucleotides comprise a ribonucleic acid domain and a blocking group at or near the 3′ end of the oligonucleotide. These compounds offer an added level of specificity previously unseen. Methods for performing nucleic acid amplification, ligation and sequencing are also provided. Additionally, kits containing the oligonucleotides are also disclosed herein.
1. A hot start enzyme composition comprising an RNase H enzyme and a chemical modification to said RNase H enzyme, wherein said chemical modification reversibly inactivates said RNase H enzyme, and wherein said chemical modification is formed by reacting said RNase H enzyme with a maleic acid anhydride analog.
2. The hot start enzyme composition of claim 1 , wherein said RNase H enzyme is Pyrococcus abyssi RNase H enzyme.
3. The hot start enzyme composition of claim 2 , wherein said Pyrococcus abyssi RNase H enzyme comprises the amino acid sequence encoded by the nucleotide sequence of SEQ ID NO: 4.
4. The hot start enzyme composition of claim 1 , wherein said maleic anhydride analog is citraconic anhydride.
5. A hot start enzyme composition comprising an RNase H enzyme and an antibody bound to said RNase H enzyme, wherein said antibody reversibly inactivates said RNase H enzyme when said antibody is bound to said RNase H enzyme.
6. The hot start enzyme composition of claim 5 , wherein said RNase H enzyme is Pyrococcus abyssi RNase H enzyme.
7. The hot start enzyme composition of claim 6 , wherein said Pyrococcus abyssi RNaseH enzyme comprises the amino acid sequence encoded by the nucleotide sequence of SEQ ID NO: 4.
8. The hot start enzyme composition of claim 6 , wherein said antibody is an antibody that specifically binds said Pyrococcus abyssi RNase H enzyme.