Linking sequence reads using paired code tags

• US 10,246,705 B2
• Filed: 05/19/2016
• Issued: 04/02/2019
• Est. Priority Date: 02/10/2011
• Status: Active Grant

• Current Assignee: Illumina Incorporated
• Original Assignee: Ilumina Incorporated
• Inventors: Steemers, Frank J., Gunderson, Kevin L., Royce, Thomas, Pignatelli, Natasha, Goryshin, Igor, Caruccio, Nicholas
• Primary Examiner(s): Dunston, Jennifer
• Application Number:
  US15/159,588
• Publication Number:
  US 20160251650A1
• Time in Patent Office: 1,048 Days
• Field of Search: None
• CPC Class Codes: C12N 15/1082 Preparation or screening gene libraries by chromosomal integration of polynucleotide sequences, HR-, site-specific-recombination, transposons, viral vectors
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: C12Q 2521/301 Endonuclease
• CPC Class Codes: C12Q 2521/319 Exonuclease
• CPC Class Codes: C12Q 2521/327 RNAse, e.g. RNAseH
• CPC Class Codes: C12Q 2521/507 Recombinase
• CPC Class Codes: C12Q 2525/155 incorporating/generating a new priming site
• CPC Class Codes: C12Q 2525/161 incorporating target specific and non-target specific sites
• CPC Class Codes: C12Q 2525/185 incorporating bases where the precise position of the bases in the nucleic acid string is important
• CPC Class Codes: C12Q 2525/186 incorporating a non-extendable or blocking moiety
• CPC Class Codes: C12Q 2525/191 incorporating an adaptor
• CPC Class Codes: C12Q 2525/197 incorporating a spacer/coupling moiety
• CPC Class Codes: C12Q 2525/313 Branched oligonucleotides
• CPC Class Codes: C12Q 2535/122 Massive parallel sequencing
• CPC Class Codes: C12Q 2563/179 the label being a nucleic acid
• 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

Abstract

Artificial transposon sequences having code tags and target nucleic acids containing such sequences. Methods for making artificial transposons and for using their properties to analyze target nucleic acids.

Claims

1. An artificial transposon, comprising:

a first transposase recognition site,

a second transposase recognition site,

a barcode disposed therebetween, wherein the barcode comprises a double-stranded nucleic acid sequence comprising

a first strand barcode and

a second strand barcode, wherein the first strand barcode and the second strand barcode comprise non-complementary sequences; and

a fragmentation site disposed within the first strand barcode and the second strand barcode.

2. The transposon of claim 1, wherein the first transposase recognition site comprises a hyperactive Tn5 transposase recognition site.

3. The transposon of claim 1, wherein the first transposase recognition site comprises a Mu transposase recognition site.

4. The transposon of claim 1, wherein the first transposase recognition site comprises an IS911 transposase recognition site.

5. The transposon of claim 1, further comprising at least one universal primer site.

6. An isolated template nucleic acid, comprising at least a portion of a target nucleic acid or copy thereof and at least a first transposon and second transposon of claim 1, wherein the barcode of the first transposon is different from the barcode of the second transposon.

7. The transposon of claim 1, wherein both the first strand barcode and the second strand barcode are at least four nucleotides in length.

8. The transposon of claim 1, wherein the fragmentation site comprises a nickase recognition site.

9. A DNA library preparation kit, comprising:

a plurality of artificial transposons, wherein each artificial transposon of the plurality of artificial transposon comprises;

a first transposase recognition site;

a second transposase recognition site;

a barcode disposed therebetween, wherein the barcode comprises a double-stranded nucleic acid sequence comprising

a first strand barcode and

a second strand barcode, wherein the first strand barcode and the second strand barcode comprise non-complementary sequences; and

a fragmentation site disposed within the first strand barcode and the second strand barcode; and

wherein each barcode of each artificial transposon of the plurality of artificial transposons is distinguishable from one another.

10. The DNA library preparation kit of claim 9, wherein both the first strand barcode and the second strand barcode are at least four nucleotides in length.