Error correction in amplification of samples

• US 10,202,641 B2
• Filed: 05/30/2017
• Issued: 02/12/2019
• Est. Priority Date: 05/31/2016
• Status: Active Grant

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Cellular Research, Inc. (Becton, Dickinson & Co)
• Inventors: Shum, Eleen
• Primary Examiner(s): Ketter, James S
• Application Number:
  US15/608,780
• Publication Number:
  US 20170342465A1
• Time in Patent Office: 623 Days
• Field of Search: None
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2521/107 RNA dependent DNA polymerase,(i.e. reverse transcriptase)
• CPC Class Codes: C12Q 2565/514 characterised by the use of the arrayed oligonucleotides as identifier tags, e.g. universal addressable array, anti-tag or tag complement array
• CPC Class Codes: C12Q 2565/519 characterised by the capture moiety being a single stranded oligonucleotide

Abstract

Disclosed herein are methods and systems for correcting errors in sample amplification, including the errors occurred in determining the number of targets in samples. In some embodiments, the method comprises: stochastically barcoding a plurality of targets in the samples using oligonucleotides comprising stochastic barcodes to generate stochastically barcoded targets; contacting one or more defined barcoded primers with each of the one or more samples; and determining an amplification noise.

Claims

1. A kit comprising:

a plurality of oligonucleotides comprising stochastic barcodes, wherein each stochastic barcode comprises a sample label and a molecular label, wherein the sample labels of at least two stochastic barcodes have different sequences, and wherein the molecular labels of stochastic barcodes with sample labels of the same sequence differ from one another by at least one nucleotide;

defined barcoded primers, wherein each defined barcoded primer comprises a defined sample label and a defined molecular label, wherein the defined sample labels of defined barcoded primers are variants of the sample labels of the stochastic barcodes, and wherein the Hamming distance between the sample label and the defined sample label is at least 2; and

instructions for using the plurality oligonucleotides comprising the plurality of oligonucleotides and the defined barcoded primers.

2. The kit of claim 1, wherein the defined sample label is 5-20 nucleotides in length.

3. The kit of claim 1, wherein the defined molecular label is 5-20 nucleotides in length.

4. The kit of claim 1, wherein the defined molecular label and the molecular label have the same length.

5. The kit claim 1, wherein different types of defined barcoded primers have the same length.

6. The kit of claim 1, wherein the lengths of different types of defined barcoded primers differ by at most 10 nucleotides.

7. The kit of claim 1, wherein the sample label and the defined sample label have the same length.

8. The kit of claim 1, wherein the Hamming distance between the sample label and the defined sample label is at least 4.

9. The kit of claim 1, wherein the stochastic barcodes and the defined barcoded primers have the same length.

10. The kit of claim 1, wherein the stochastic barcodes and the defined barcoded primers have different lengths.

11. The kit of claim 1, wherein the kit comprises a synthetic particle associated with the plurality of oligonucleotides comprising the stochastic barcodes.

12. The kit of claim 11, wherein the kit comprises a microwell pre-loaded with the synthetic particle.

13. The kit of claim 11, wherein the synthetic particle is a bead.

14. The kit of claim 13, wherein the bead is a hydrogel bead.

15. The kit of claim 13, wherein the bead is a magnetic bead.

16. The kit of claim 11, wherein the synthetic particle is a silica gel bead, a controlled pore glass bead, a Dynabead, a Sephadex/Sepharose bead, a cellulose bead, a polystyrene bead, or a combination thereof.

17. The kit of claim 1, comprising a buffer.

18. The kit of claim 1, comprising a cartridge.

19. The kit of claim 1, comprising one or more reagents for a reverse transcription reaction.

20. The kit of claim 1, comprising one or more reagents for an amplification reaction.