Patent US9976181

Synthetic nucleic acid spike-ins
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US9976181

1. A method for determining abundance of nucleic acids in an initial sample comprising target nucleic acids, the method comprising: (19) (3)

(a) adding a starting quantity of at least 1,000 unique synthetic nucleic acids to the initial sample, wherein each of the at least 1,000 unique synthetic nucleic acids comprises (i) an identifying tag and (ii) a variable region comprising at least 5 degenerate bases;

(b) performing a sequencing assay on a portion of the target nucleic acids and on a portion of the at least 1,000 unique synthetic nucleic acids, thereby obtaining target and synthetic nucleic acid sequence reads; and

(c) calculating a diversity loss value for the portion of the at least 1,000 unique synthetic nucleic acids, wherein abundance of the target nucleic acids in the initial sample is determined using the diversity loss value.

2. The method of claim 1, wherein the target nucleic acids comprise pathogen nucleic acids. (1) (0)

4. The method of claim 1, wherein the at least 1,000 unique synthetic nucleic acids comprise DNA. (0)

5. The method of claim 1, wherein each of the at least 1,000 unique synthetic nucleic acids is less than 500 base pairs or nucleotides in length. (0)

6. The method of claim 1, wherein the initial sample is blood, plasma, serum, cerebrospinal fluid, synovial fluid, bronchial-alveolar lavage, urine, stool, saliva, or a nasal sample. (0)

7. The method of claim 1, wherein the initial sample is a sample of isolated nucleic acids. (0)

8. The method of claim 1, further comprising generating a sequencing library from the initial sample, wherein the at least 1,000 unique synthetic nucleic acids are added to the sample before the generating of the sequencing library. (0)

9. The method of claim 1, wherein the diversity loss value for the portion of the at least 1,000 unique synthetic nucleic acids indicates a loss of one or more nucleic acids during sample processing of the initial sample. (0)

10. The method of claim 1, wherein the identifying tag of each of the at least 1,000 unique synthetic nucleic acids comprises a common sequence. (0)

11. The method of claim 1, wherein the initial sample is from a human subject. (0)

12. The method of claim 1, wherein the at least 1,000 unique synthetic nucleic acids comprise at least 10⁴unique synthetic nucleic acids. (0)

13. The method of claim 1, further comprising adding additional synthetic nucleic acids to the initial sample, wherein the additional synthetic nucleic acids have at least three different lengths. (2) (0)

14. The method of claim 1, further comprising adding to the initial sample a first group of additional synthetic nucleic acids with a first length, a second group of additional synthetic nucleic acids with a second length, and a third group of additional synthetic nucleic acids with a third length, wherein the first group of additional synthetic nucleic acids, the second group of additional synthetic nucleic acids and third group of additional synthetic nucleic acids each comprises synthetic nucleic acids with at least three different GC contents. (0)

17. The method of claim 1, wherein the at least 1,000 unique synthetic nucleic acids are added to the initial sample at a first sample processing step. (1) (0)

18. The method of claim 17, further comprising adding an additional pool of at least 1,000 unique synthetic nucleic acids to the initial sample at a second sample processing step, wherein the second sample processing step is different from the first sample processing step. (3) (0)

22. The method of claim 1, further comprising adding a sample identifier nucleic acid to the initial sample. (0)

23. The method of claim 1, wherein (a) further comprises adding non-unique synthetic nucleic acids to the initial sample. (0)

24. The method of claim 1, further comprising: reporting a result of the method to a caregiver, patient, or other person. (0)

25. The method of claim 1, wherein the determined abundance is a relative abundance. (0)

26. The method of claim 1, wherein the determined abundance is absolute abundance. (0)

27. The method of claim 1, further comprising determining a number of synthetic nucleic acid sequence reads comprising different variable regions. (2) (0)

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Patent US9976181

Synthetic nucleic acid spike-ins

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