Methods of specifically labeling nucleic acids using CRISPR/Cas
US11761028B2 · kind B2 · utility
Assignee
Inventors
Key dates
| Filing date | Oct 18, 2017 |
| Grant date | Sep 19, 2023 |
| Priority date | — |
| Expiry date | May 10, 2040 |
Classification
- Technology area (CPC C)Chemistry; Metallurgy
- CPC primaryC12Q2563/107
- WIPO fieldBiotechnology
- WIPO sectorChemistry
Abstract
A method of detecting the length of an individual telomere is provided. In one embodiment, the method includes contacting genomic DNA with a guide RNA having a portion complementary to a telomere repeat sequence in the genomic DNA and with Cas9 nickase to produce a single-strand break in the genomic DNA at the telomere repeat sequence. The nicked DNA is contacted with a polymerase and fluorescently labeled nucleotide, wherein the fluorescently labeled nucleotide is incorporated into the nicked DNA at the telomere repeat sequence. The genomic DNA is contacted with a second nicking endonuclease which is specific for a sequence motif in the genomic DNA thereby producing a second nick in the genomic DNA at the motif sequence. The nicked DNA is contacted with a polymerase and second fluorescently labeled nucleotide of different color, wherein the second fluorescently labeled nucleotide is incorporated into the nicked DNA at the motif sequence location. The length of the telomere is detected by measuring the fluorescence of first fluorescently labeled nucleotide at the telomere repeat location, wherein the fluorescently labeled motif sequences are used as a barcode to identify the chromo…
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.