RARE CELL ANALYSIS USING SAMPLE SPLITTING AND DNA TAGS

• US 20080220422A1
• Filed: 06/14/2007
• Published: 09/11/2008
• Est. Priority Date: 06/14/2006
• Status: Active Grant

• Current Assignee: Verinata Health Incorporated (Illumina Incorporated), Mehmet Toner, ZEON Corporation
• Original Assignee: Mehmet Toner
• Inventors: Toner, Mehmet, Kapur, Ravi, Wang, Zihua, Gray, Darren, Krueger, Neil X., Fuchs, Martin, Shoemaker, Daniel
• Granted Patent: US 8,372,584 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: C12Q 1/6809 Methods for determination or identification of nucleic acids involving differential detection
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6881 for tissue or cell typing, e.g. human leukocyte antigen [HLA] probes
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: C12Q 2600/16 Primer sets for multiplex assays
• CPC Class Codes: G01N 2015/1006 for cytology
• CPC Class Codes: G01N 2015/1029 Particle size
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

The present invention provides systems, apparatuses, and methods to detect the presence of fetal cells when mixed with a population of maternal cells in a sample and to test fetal abnormalities, e.g. aneuploidy. The present invention involves labeling regions of genomic DNA in each cell in said mixed sample with different labels wherein each label is specific to each cell and quantifying the labeled regions of genomic DNA from each cell in the mixed sample. More particularly the invention involves quantifying labeled DNA polymorphisms from each cell in the mixed sample.

Claims

1. A method for fetal diagnosis comprising:

labeling one or more genomic DNA regions in fetal and non-fetal cells, enriched from a maternal blood sample, using labels adapted to distinguish between individual cells, and determining the presence or absence of a fetal abnormality by analyzing said labeled genomic DNA regions.

3. The method of claim 1, further comprising splitting said fetal and non-fetal cells such that individual cells are in addressable locations.

5. The method of claim 3, further comprising amplifying said one or more genomic DNA regions from each cell to generate a plurality of amplicons.

6. The method of claim 5, further comprising pooling said amplicons.

7. The method of claim 5 wherein said regions of genomic DNA comprise one or more polymorphisms.

8. The method of claim 7, wherein said polymorphisms are STRs or SNPs.

11. The method of claim 3, wherein said labels each comprise a unique tag sequence specific for each addressable location.

12. The method of claim 1 wherein said label comprises a nucleic acid sequence between 4-20 bp long that is unique to each cell

13. The method of claim 1, wherein at least two genomic DNA regions on a chromosome of interest are labeled.

14. The method of claim 1, wherein said genomic DNA regions are located on two or more chromosomes of interest.

15. The method of claim 1, wherein said analyzing comprises performing ultra deep sequencing or quantitative genotyping.

16. The method of claim 1, wherein said fetal abnormality comprises an aneuploidy selected from the group consisting of:

trisomy 13, trisomy 18, trisomy 21 (Down Syndrome), Klinefelter Syndrome (XXY), monosomy, triploidy, tetraploidy, or other irregular number of sex or autosomal chromosomes, and a combination thereof.

17. The method of claim 1, wherein said fetal abnormality comprises an aneuploidy that is maternally or paternally derived.

18. The method of claim 1, wherein the fetal abnormality is a segmental aneuploidy.

46. The method of claim 1, further comprising binning at least a portion of said enriched cells

48. The method of claim 46, wherein said binning is performed using serial dilution.

2. A method for fetal diagnosis comprising:

labeling one or more cDNA sequences generated from fetal and non-fetal cells, wherein said fetal and non-fetal cells are enriched from a maternal blood sample, using labels adapted to distinguish between individual cells, and

determining the presence or absence of a fetal abnormality by analyzing said labeled cDNA regions.

4. The method of claim 2, further comprising splitting said fetal and non-fetal cells such that individual cells are in addressable locations.

9-10. -10. (canceled)

19. (canceled)

20. A method for fetal diagnosis comprising:

amplifying a plurality of genomic DNA regions from one or more fetal cells enriched from maternal blood;

wherein the resulting amplicons comprise a cell specific label, and determining the presence or absence of a fetal abnormality by analyzing said amplicons.

22. The method of claim 20, further comprising splitting said one or more fetal cells such that each individual cell is in an addressable location.

28. The method of claim 20, wherein said labels each comprise a unique tag sequence specific for each addressable location.

21. A method for fetal diagnosis comprising:

amplifying a plurality of cDNA sequence generated from one or more fetal cells enriched from maternal blood;

wherein the resulting amplicons comprise a cell specific label, and determining the presence or absence of a fetal abnormality by analyzing said amplicons.

23-27. -27. (canceled)

29-36. -36. (canceled)

37. Amended) A computer readable medium comprising logic for analyzing a plurality of amplicons, from one or more genomic DNA regions on one or more chromosomes, or one or more cDNA sequences from at least one fetal cell and at least one maternal cell, enriched from maternal blood.

39. The computer readable medium of claim 37, wherein said logic analyzes the ratio of maternal to fetal chromosome copy number.

40. The method of claim 37, wherein said amplicons comprise a unique tag sequence

38. (canceled)

41-45. -45. (canceled)

47. (canceled)

49. (canceled)

50-54. -54. (canceled)