Enrichment of nucleic acid targets
Methods and apparatus providing improved fidelity and specificity when separating nucleic acids from a sample, but without need for amplification. In particular, using the disclosed methods, it is possible to isolate a variant nucleic acid (i.e., a mutation) from a non-target nucleic acid (i.e., a wild-type) when the variant is present in the original sample at a much lower concentration than the non-target, e.g., 1:10,000, without substantial loss of the variant.
1. A method for enriching a sample, the method comprising the steps of:
obtaining a sample comprising a non-target nucleic acid and a target nucleic acid, wherein the target nucleic acid and the non-target nucleic acid differ by only a single base at a position of interest but do not differ in length, and the target to non-target ratio in the sample is X; and
enriching for the target nucleic acid by applying a time-varying driving field to the sample in the presence of a separation medium,
wherein:
(a) after enrichment the ratio of target nucleic acid to non-target nucleic acid is Y, wherein Y is at least 10 times greater than X, and
no new molecules are generated in the sample during said enriching step.
2. The method of claim 1 , wherein the target and the non-target have different methylation or acetylation patterns.
3. The method of claim 1 , wherein the sample is a biological sample.
4. The method of claim 3 , wherein the biological sample comprises tissue, blood, sputum, sweat, urine, feces, tears, aspirate, or a combination thereof.
5. The method of claim 4 , wherein the biologic sample is from a human.
6. The method of claim 1 , further comprising identifying the target nucleic acid.
7. The method of claim 6 , wherein identifying comprises sequencing at least a portion of the target nucleic acid, contacting the target nucleic acid with a labeled moiety, or an amplification-based detection.
8. The method of claim 1 , wherein the presence of the target nucleic is indicative of a disorder.
9. The method of claim 8 , wherein the disorder is a genetic disorder.
10. The method of claim 8 , wherein the disorder is cancer.
11. The method of claim 1 , wherein Y is at least 100 times greater than X.
12. The method of claim 11 , wherein Y is at least 1,000 times greater than X.
13. The method of claim 1 , wherein enriching comprises applying a periodic field to the sample.
14. The method of claim 13 , wherein the periodic field is a periodic electric field.
15. The method of claim 1 , wherein enriching comprises applying two periodic fields to the sample.
16. The method of claim 15 , wherein one periodic field is electric and the other is thermal.
17. The method of claim 1 , wherein the non-target and target nucleic acids each comprise 1000 or fewer bases.
18. The method of claim 17 , wherein the non-target and target nucleic acids each comprise 200 or fewer bases.
19. The method of claim 18 , wherein the no target and target nucleic acids each comprise 30 or fewer bases.
20. The method of claim 1 , wherein the non-target nucleic acid is a wild-type nucleic acid.
21. The method of claim 1 , wherein at least 20% of the target in the sample comprising a non-target nucleic acid and a target nucleic acid is recovered after enrichment.
22. The method of claim 1 , wherein the sample is simultaneously enriched for more than one target nucleic acid.
23. The method of claim 22 , wherein the presence of more than one target nucleic acid is indicative of a disorder.
24. The method of claim 22 , wherein the sample is simultaneously enriched for more than twenty target nucleic acids.
25. The method of claim 24 , wherein the sample is simultaneous , enriched for more than 100 target nucleic acids.
26. The method of claim 1 , wherein the sample comprises about 1% or less target nucleic acid (mol target nucleic acid/mol non-target nucleic acid).