US2020251180A1PendingUtilityA1
Resolving genome fractions using polymorphism counts
Est. expiryApr 12, 2031(~4.7 yrs left)· nominal 20-yr term from priority
G16B 40/00G16B 30/10G16B 20/10G16B 20/20G16B 30/00C12Q 1/6809C12Q 1/6883C12Q 2600/156C12Q 1/6876G16B 45/00G16B 20/00C12Q 1/6827
73
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Claims
Abstract
Methods of reliably estimating genomic fraction (e.g., fetal fraction) from polymorphisms such as small base variations or insertions-deletions are disclosed. Sequenced data from a multigenomic source is used to determine allele counts for one or more of the polymorphisms. For one or more of the polymorphisms, zygosity is assigned, and genomic fraction is determined from the zygosity and allele counts. Certain embodiments employ SNPs as the relevant polymorphism. The disclosed methods can be applied as part of an intentional, pre-designed re-sequencing study targeted against known polymorphisms or can be used in a retrospective analysis of variations found by coincidence in overlapping sequences generated from maternal plasma (or any other setting where a mixture of DNA from several people are present).
Claims
exact text as granted — not AI-modified1 . A method, implemented at a computer system that includes one or more processors and system memory, of estimating a fraction of fetal DNA in DNA obtained from a bodily fluid of a pregnant individual carrying a fetus, the method comprising:
(a) sequencing DNA obtained from the bodily fluid of the pregnant individual using a nucleic acid sequencer under conditions that produce DNA sequence reads containing a plurality of polymorphisms; (b) mapping, by the computer system, the DNA sequence reads to a plurality of polymorphism sites on a reference sequence corresponding to the plurality of polymorphisms; (c) determining, by the computer system, allele frequencies of the DNA sequence reads for the plurality of polymorphisms; and (d) applying, by the computer system, the allele frequencies to a mixture model to estimate the fraction of fetal DNA in the DNA obtained from the bodily fluid of the individual carrying the fetus.
2 . The method of claim 1 , wherein the mixture model employs factorial moments to calculate the fraction of fetal DNA.
3 . The method of claim 2 , wherein the factorial moments are calculated as:
F
1
=
1
n
∑
i
=
1
n
a
i
d
i
F
2
=
1
n
∑
i
=
1
n
a
i
(
a
i
-
1
)
d
i
(
d
i
-
1
)
…
F
j
=
1
n
∑
i
=
1
n
a
i
(
a
i
-
1
)
…
(
a
i
-
j
+
1
)
d
i
(
d
i
-
1
)
(
d
i
-
j
+
1
)
wherein F j is the j th factorial moment, a i is a minor allele frequency of the i th polymorphism, d i is a coverage of the i th polymorphism, and n is the number of polymorphisms.
4 . The method of claim 3 , wherein (d) comprises estimating the fraction of fetal DNA based on the following relations between the factorial moments and probabilities of minor allele in zygosity cases:
F
1
≈
∑
i
=
1
m
α
i
p
i
1
F
2
≈
∑
i
=
1
m
α
i
p
i
2
…
F
j
≈
∑
i
=
1
m
α
i
p
i
j
…
F
g
≈
∑
i
=
1
m
α
i
p
i
g
wherein α i is the portion of the plurality of polymorphisms in zygosity case i, p i is a binomial probability of minor allele in zygosity case i, m is the number of zygosity cases, and g is the number of calculated factorial moments;
and wherein the zygosity cases are selected from: (i) the pregnant individual is homozygous and the fetus is homozygous, (ii) the pregnant individual is homozygous and the fetus is heterozygous, (iii) the pregnant individual is heterozygous and the fetus is homozygous, and (iv) the pregnant individual is heterozygous and the fetus is heterozygous.
5 . The method of claim 4 , wherein the plurality of polymorphisms consists of polymorphisms in cases (i) and (ii), the method further comprising calculating the fraction of fetal DNA based on the following relations:
F
1
≈
α
1
e
+
(
1
-
α
1
)
(
f
2
)
F
2
≈
α
1
e
2
+
(
1
-
α
1
)
(
f
2
)
2
F
3
≈
α
1
e
3
+
(
1
-
α
1
)
(
f
2
)
3
wherein F 1 , F 2 , and F 3 are the first three factorial moments, α 1 is the portion of the plurality of polymorphisms in zygosity case (i), e is an unknown sequencing error rate, and f is the fraction of fetal DNA.
6 . The method of claim 4 , wherein the plurality of polymorphisms consists of polymorphisms in cases (i) and (ii), the method further comprising calculating the fraction of fetal DNA based on the following relations:
F
1
≈
α
1
e
+
(
1
-
α
1
)
(
f
2
)
F
2
≈
α
1
e
2
+
(
1
-
α
1
)
(
f
2
)
2
wherein F 1 and F 2 are the first two factorial moments, α 1 is the portion of the plurality of polymorphisms in zygosity case (i), e is a known sequencing error rate, and f is the fraction of fetal DNA.
7 . The method of claim 4 , wherein the plurality of polymorphisms consists of polymorphisms in cases (i) and (ii), the method further comprising calculating the fraction of fetal DNA based on the following relations:
F
1
=
α
1
e
4
+
(
1
-
α
1
)
(
e
+
f
2
)
F
2
=
α
1
(
e
4
)
2
+
(
1
-
α
1
)
(
e
+
f
2
)
2
wherein F 1 and F 2 are the first two factorial moments, α 1 is the portion of the plurality of polymorphisms in zygosity case (i), e is a sequencing error rate that is differentially applied to cases (i) and (ii), and f is the fraction of fetal DNA.
8 . The method of claim 1 , further comprising, prior to (d), computationally removing allele frequencies for polymorphisms identified as being heterozygous in both the fetus and the pregnant individual.
9 . The method of claim 1 , further comprising, prior to (d), computationally removing allele frequencies for polymorphisms identified as being homozygous in both the fetus and the pregnant individual.
10 . The method of claim 1 , further comprising, prior to (d), computationally removing allele frequencies for polymorphisms identified as being heterozygous in the pregnant individual.
11 . The method of claim 1 , wherein the mixture model accounts for sequencing error.
12 . The method of claim 1 , wherein the DNA obtained from a bodily fluid of a pregnant individual comprises cell-free DNA obtained from a plasma sample of the pregnant individual.
13 . The method of claim 1 , further comprising obtaining a sample of the bodily fluid of the pregnant individual.
14 . The method of claim 1 , wherein the mapping in (b) comprises identifying a plurality of biallelic polymorphism sequences.
15 . The method of claim 1 , further comprising, based on the estimate of the fraction of fetal DNA, prescribing, initiating, and/or altering treatment of a human subject from whom the bodily fluid of the pregnant individual was taken.
16 . The method of claim 1 , further comprising, based on the estimate of the fraction of fetal DNA, ordering and/or performing one or more additional tests.
17 . The method of claim 1 , wherein the sequencing is conducted without selectively amplifying any of the plurality of polymorphisms.
18 . An apparatus for estimating a fraction of fetal DNA in DNA obtained from a bodily fluid of a pregnant individual carrying a fetus, the apparatus comprising system memory and one or more processors configured to:
sequence DNA extracted from a sample of the bodily fluid comprising DNA of both a maternal genome and a fetal genome to produce DNA sequence reads containing a plurality of polymorphisms; map the DNA sequence reads to a plurality of polymorphism sites on a reference sequence corresponding to the plurality of polymorphisms; determine allele frequencies of the mapped DNA sequence reads for the plurality of polymorphisms; and apply the allele frequencies to a mixture model to estimate the fraction of fetal DNA in the DNA obtained from the bodily fluid of the individual carrying the fetus.
19 . The apparatus of claim 18 , wherein the mixture model employs factorial moments to calculate the fraction of fetal DNA.
20 - 28 . (canceled)
29 . A computer program product comprising a non-transitory machine readable medium storing program code that, when executed by one or more processors of a computer system, causes the computer system to implement a method of estimating a fraction of fetal DNA in DNA obtained from a bodily fluid of a pregnant individual carrying a fetus, said program code comprising:
(a) code for sequencing DNA obtained from the bodily fluid of the pregnant individual using a nucleic acid sequencer under conditions that produce DNA sequence reads containing a plurality of polymorphisms; (b) code for mapping the DNA sequence reads to a plurality of polymorphism sites on a reference sequence corresponding to the plurality of polymorphisms; (c) code for determining allele frequencies of the DNA sequence reads for the plurality of polymorphisms; and (d) code for applying the allele frequencies to a mixture model to estimate the fraction of fetal DNA in the DNA obtained from the bodily fluid of the individual carrying the fetus, wherein the mixture model employs factorial moments to calculate the fraction of fetal DNA.
30 - 40 . (canceled)Join the waitlist — get patent alerts
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