Non-invasive fetal genetic screening by digital analysis
AssigneeThe Board of Trustees of the Leland Stanford Junior University
InventorsStephen Quake, Hei-Mun Christina Fan
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Abstract
The present methods are exemplified by a process in which maternal blood containing fetal DNA is diluted to a nominal value of approximately 0.5 genome equivalent of DNA per reaction sample. Digital analysis is then be used to detect aneuploidy, such as the trisomy that causes Down Syndrome. Since aneuploidies do not present a mutational change in sequence, and are merely a change in the number of chromosomes, it has not been possible to detect them in a fetus without resorting to invasive techniques such as amniocentesis or chorionic villi sampling. Digital amplification allows the detection of aneuploidy using massively parallel amplification and detection methods, examining, e.g., 10,000 genome equivalents.
Claims14
A method for identifying a risk for an aneuploidy on a first chromosome of a fetus carried by a euploid mother, the method comprising:
- a) distributing a sample of nucleic acids into at least 500 discrete locations, wherein said sample is derived from a cell-free portion of a maternal blood sample that contains a mixture of maternal and fetal DNA;
- b) amplifying said nucleic acids at said discrete locations, wherein each of said discrete locations contains, on average, not more than about one target sequence per discrete location;
- c) determining a first value by counting a number of detected sequences at the at least 500 locations representing a first location on the first chromosome, wherein counting comprises detecting fluorescently-labeled probes hybridized to the amplified nucleic acids at said discrete locations, wherein the first value:
- i) includes a contribution from detection of the maternal DNA and the fetal DNA, and
- ii) is not based on separately determining contributions to the amount at the first location from the maternal DNA and the fetal DNA;
- d) determining a second value by counting a number of detected sequences at the at least 500 locations representing a second location on the second chromosome which is presumed to be euploid, wherein counting comprises detecting fluorescently-labeled probes hybridized to the amplified nucleic acids at said discrete locations, wherein the second value:
- i) includes a contribution from detection of the maternal DNA and the fetal DNA;
- and
- ii) is not based on separately determining contributions to the amount at the second location from the maternal DNA and the fetal DNA;
- and
- e) determining a difference between the first value and the second value, wherein a statistically significant difference indicates the presence of aneuploidy on the first chromosome in the fetal DNA.
A method for identifying a risk for an aneuploidy on a first chromosome of a fetus carried by a euploid mother, the method comprising:
- a) distributing a sample of nucleic acids into at least 500 discrete locations, wherein said sample is derived from a cell-free portion of a maternal blood sample that contains a mixture of maternal and fetal DNA;
- b) amplifying said nucleic acids at said discrete locations, wherein each of said discrete locations contains, on average, not more than about one target sequence per discrete location;
- c) determining first multiple values by counting a number of detected sequences at multiple different locations on the first chromosome at the at least 500 locations, wherein counting comprises detecting fluorescently-labeled probes hybridized to the amplified nucleic acids at said discrete locations, wherein the first multiple values:
- i) include a contribution from detection of the maternal DNA and the fetal DNA;
- and
- ii) are not based on separately determining contributions to the amounts at the multiple different locations on the first chromosome from the maternal DNA and the fetal DNA;
- and
- d) determining second multiple values by counting a number of detected sequences at multiple different locations on the second chromosome which is presumed to be euploid at the at least 500 locations, wherein counting comprises detecting fluorescently-labeled probes hybridized to the amplified nucleic acids at said discrete locations, wherein the second multiple values:
- i) include a contribution from detection of the maternal DNA and the fetal DNA;
- and
- ii) are not based on separately determining contributions to the amounts at the multiple different locations on the second chromosome from the maternal DNA and the fetal DNA;
- and
- e) determining a difference between the first multiple values, and the second multiple values wherein a statistically significant difference indicates the presence of aneuploidy on the first chromosome in the fetal DNA.
Dependencies are inferred from the claim text.