Process for controlling contamination of nucleic acid amplification reactions
View Patent ↗This invention relates to a method of incorporating an exo-sample nucleotide into the amplified product strands resulting from a nucleic acid amplification process. Once the product strands have been obtained and analyzed (e.g., by hybridization, Southern blot, etc.), the exo-sample strands can be selectively destroyed by acting on the incorporated exo-sample nucleotide. Two embodiments are presented. In a first embodiment, the exo-sample nucleotide is incorporated by carrying out the amplification to reaction in the presence of an excess of exo-sample nucleotide tri-phosphate. In a second embodiment, the exo-sample nucleotide is incorporated by carrying out the amplification reaction in the presence of an oligonucleotide which has, as part of its sequence, one or more exo-sample nucleotides.
1. A reaction mixture that incorporates dUTP into a nucleic acid amplification product, said mixture comprising a thermostable DNA polymerase, a detergent, dUTP, and a deoxyuridine-containing nucleic acid, wherein said nucleic acid is double-stranded and has deoxyuridine in one strand.
2. The reaction mixture of claim 1 , wherein said reaction mixture is a polymerase chain reaction (PCR).
3. The reaction mixture of claim 1 , wherein said thermostable DNA polymerase is Taq DNA polymerase.
4. The reaction mixture of claim of claim 1 , further comprising a uracil DNA glycosylase.
5. A DNA amplification composition comprising a thermostable DNA polymerase, a detergent, exo-sample nucleotides, and a double-stranded nucleic acid comprising: (a) a first nucleic acid strand comprising no exo-sample nucleotides, and (b) a second nucleic acid strand comprising one or more exo-sample nucleotides.
6. The composition of claim 5 , wherein the second nucleic acid strand further comprises a primer joined to an extension product of the primer, and wherein the extension product of the primer comprises one or more exo-sample nucleotides.
7. A DNA amplification composition comprising a thermostable DNA polymerase, a detergent, dUTP, and a double-stranded nucleic acid comprising: (a) a first nucleic acid strand comprising a sugar-phosphate backbone having no apyrimidinic deoxyriboses, and (b) a second nucleic acid strand comprising a sugar-phosphate backbone having one or more apyrimidinic deoxyriboses.
8. The composition of claim 7 , wherein the second nucleic acid strand further comprises a primer joined to an extension product of the primer, and wherein the extension product of the primer comprises one or more apyrimidinic deoxyriboses.
9. The composition of claim 5 , wherein said composition is a polymerase chain reaction (PCR).
10. The composition of claim 5 , wherein said thermostable DNA polymerase is Taq DNA polymerase.
11. The composition of claim 5 , further comprising a uracil DNA glycoslase.
12. The composition of claim 7 , wherein said composition is a polymerase chain reaction (PCR).
13. The composition of claim 7 , wherein said thermostable DNA polymerase is Taq DNA polymerase.
14. The composition of claim 7 , further comprising a uracil DNA glycoslase.
15. The reaction mixture of claim 1 , further comprising double-stranded nucleic acids having deoxyuridine in both strands.
16. The composition of claim 5 , further comprising double-stranded nucleic acids having deoxyuridine in both strands.
17. The composition of claim 7 , further comprising double-stranded nucleic acids having apyrimidinic deoxyriboses in both strands.
18. The composition of claim 1 , wherein the deoxyuridine-containing strand further comprises a primer joined to an extension product of the primer, and wherein the extension product of the primer comprises deoxyuridine.