Granted Patent B2

US 12,221,653 · App. 17/685,730 · Granted Feb 11, 2025

Inventors: Joshua Babiarz (Castro Valley, CA); Tudor Pompiliu Constantin (Berkeley, CA); Lane A. Eubank (San Carlos, CA); George Gemelos (Portland, OR); Matthew Micah Hill (Belmont, CA); Huseyin Eser Kirkizlar (Los Angeles, CA); Matthew Rabinowitz (San Francisco, CA); Onur Sakarya (Redwood City, CA); Styrmir Sigurjonsson (San Jose, CA); Bernhard Zimmermann (Manteca, CA)

Assignee: Natera, Inc.

C12Q1/6883C12Q1/6809C12Q1/6811C12Q1/6844C12Q1/6848C12Q1/6851C12Q1/6855C12Q1/6869C12Q1/6874C12Q2600/156

Abstract

Claims (13)

1. A method for preparing a non-naturally occurring preparation of amplified DNA from a maternal blood sample or a fraction thereof useful for non-invasive prenatal aneuploidy determination, wherein the maternal blood sample is from a pregnant mother of a fetus, the method comprising:

(a) extracting cell-free DNA from the maternal blood sample or fraction thereof, wherein the extracted cell-free DNA comprises a mixture of fetal cell-free DNA and maternal cell-free DNA;

(b) preparing a non-naturally occurring preparation of amplified DNA by (i) hybridizing a plurality of circularizing probes to the extracted mixture of fetal and maternal cell-free DNA, wherein the circularizing probes bind to a plurality of target loci on one or more chromosomes of interest, and wherein the plurality of circularizing probes comprise at least 50 circularizing probes each targeting a different target loci, (ii) obtaining a plurality of circularized probes by ligation, wherein the circularized probes comprise sequences of the target loci, and (iii) amplifying the circularized probes by rolling-circle amplification; and

(c) analyzing the non-naturally occurring preparation of amplified DNA to measure the amounts of the target loci on the one or more chromosomes of interest from the fetal and maternal cell-free DNA.

2. The method of claim 1 , wherein one or more of the target loci map to chromosome(s) X, Y, 13, 18, and/or 21, and the method further comprises determining a ploidy state for chromosome(s) X, Y, 13, 18, and/or 21 of the fetus using the measured amounts of the target loci.

3. The method of claim 1 , wherein one or more of the target loci map to one or more reference chromosomes expected to be disomic.

4. The method of claim 1 , wherein one or more of the target loci map to chromosome(s) X, Y, 13, 18, and/or 21, and one or more of the target loci map to one or more reference chromosomes expected to be disomic.

5. The method of claim 1 , wherein the preparing step further comprises digesting the extracted cell-free DNA with a restriction enzyme.

6. The method of claim 1 , wherein the preparing step further comprises removing non-circularized probes with an exonuclease.

7. The method of claim 1 , wherein the analyzing step comprises measuring the amounts of the target loci by genotyping array, by measurement of fluorescently-tagged DNA, or by high-throughput sequencing.

8. The method of claim 1 , wherein the method further comprises determining a ploidy state of one or more chromosomes of interest of the fetus using the measured amounts of the target loci.

9. The method of claim 8 , wherein the determining a ploidy state of one or more chromosomes of interest of the fetus comprises counting the amounts of the target loci that map to a given chromosome.

10. The method of claim 8 , wherein a Z-score is determined using the amounts of the target loci on a chromosome of interest and a reference chromosome, and the Z-score is used to determine the ploidy state of the chromosome of interest.

Assignments (1)

Continuity (37)

References Cited (400)

Cited By (1)

US 12,423,216