Isolation of nucleic acids
Provided herein is technology relating to isolating nucleic acids. In particular, the technology relates to methods and kits for extracting nucleic acids from problematic samples such as stool.
1. A system for isolating a human target DNA from a stool sample from a human, the system comprising:
a) an inhibitor-adsorbing composition comprising insoluble polyvinylpolypyrrolidone particles;
b) a target-specific capture reagent comprising a solid support attached to a first oligonucleotide, wherein the first oligonucleotide is complementary to at least a portion of the human target DNA; and
c) second and third oligonucleotides, wherein the second and third oligonucleotides are a primer pair for amplifying a region of the human target DNA capturable by the target-specific capture reagent.
2. The system of claim 1 , wherein the insoluble polyvinylpolypyrrolidone particles are in pre-measured form.
3. The system of claim 2 , wherein the pre-measured form comprises a tablet.
4. The system of claim 1 , wherein the solid support is a bead or particle.
5. The system of claim 4 , wherein the bead or particle is magnetic.
6. The system of claim 5 , further comprising a magnet.
7. The system of claim 1 , wherein the first oligonucleotide is covalently attached to the solid support.
8. The system of claim 1 , further comprising a chaotropic agent.
9. The system of claim 8 , wherein the chaotropic agent comprises guanidine thiocyanate.
10. The system of claim 1 , further comprising a filter.
11. The system of claim 10 , wherein the filter is a spin filter.
12. The system of claim 1 , further comprising an elution solution.
13. The system of claim 1 , further comprising a human stool sample.
14. The system of claim 13 , wherein the human stool sample is an homogenized human stool sample comprising a buffer.
15. The system of claim 1 , further comprising a human stool supernatant collected from an homogenized human stool sample comprising a buffer.
16. The system of claim 1 , further comprising reagents for a nucleic acid detection reaction.
17. The system of claim 16 , wherein the reagents for a nucleic acid detection reaction comprise at least one enzyme selected from the group consisting of a restriction endonuclease, a flap endonuclease, a reverse transcriptase, a ligase, an RNA polymerase, and a DNA polymerase.
18. The system of claim 1 , wherein the components in a) and b) are packaged together in a kit.
19. The system of claim 1 , further comprising a detection probe.