Detection of human .alpha.-thalassemia mutations and their use as predictors of blood-related disorders
First Claim
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1. A method of screening a human subject for an increased risk of developing a blood-related disorder, comprising the steps of:
- (a) assaying genomic DNA of a human subject to determine a presence or an absence of an α
-globin deletion mutation; and
(b) screening for an increased risk of developing a blood-related disorder from the presence or absence of an α
-globin deletion mutation in said genomic DNA, wherein the presence of an α
-globin deletion mutation in the genomic DNA correlates with an increased risk of developing a blood-related disorder, and wherein said blood-related disorder is selected from the group consisting of hypertension, myocardial infarction, and diabetes.
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Abstract
The invention is based on the discovery that adults having a genotype comprising a hemoglobin α-gene deletion are significantly more likely to be hypertensive than adults having a normal (αα/αα) gentoype. The invention provides an improved method for determining a human subject'"'"'s genotype at the α-gene loci; a method of screening a human subject for an increased potential of developing hypertension and other blood-related disorders; and provides an apparatus/kit for screening a human subject for a risk of developing hypertension and other blood-related disorders.
74 Citations
39 Claims
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1. A method of screening a human subject for an increased risk of developing a blood-related disorder, comprising the steps of:
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(a) assaying genomic DNA of a human subject to determine a presence or an absence of an α
-globin deletion mutation; and(b) screening for an increased risk of developing a blood-related disorder from the presence or absence of an α
-globin deletion mutation in said genomic DNA, wherein the presence of an α
-globin deletion mutation in the genomic DNA correlates with an increased risk of developing a blood-related disorder, and wherein said blood-related disorder is selected from the group consisting of hypertension, myocardial infarction, and diabetes. - View Dependent Claims (2, 3, 4, 5, 6)
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7. A method of screening for an increased potential for developing hypertension in a normotensive human individual comprising the steps of:
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(a) isolating DNA from said human individual; (b) assaying said DNA for the presence of a deletion relative to DNA of a normal human subject, said deletion comprising a reduction in the number of α
-globin genes in the genome of the normotensive human individual; and(c) screening for an increased potential for developing hypertension in said normotensive human individual, wherein the presence of said deletion in the DNA isolated from the normotensive human individual is indicative of an increased potential for developing hypertension. - View Dependent Claims (8, 9, 10, 11, 12, 13, 14)
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15. A method for determining an increased potential for developing hypertension in a human individual comprising the steps of:
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(a) isolating genomic DNA from said human individual; (b) performing a multiplex polymerase chain reaction with said genomic DNA to amplify a marker portion and a control portion thereof, wherein the presence of an α
-globin gene deletion in said genomic DNA reduces the quantity of marker amplicons produced in the multiplex polymerase chain reaction relative to the quantity of control amplicons produced in said multiplex polymerase chain reaction; and(c) determining a potential for developing hypertension from the quantity of marker amplicons relative to the quantity of control amplicons. - View Dependent Claims (16, 17, 18, 19, 20, 21, 22)
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23. A method of determining an increased potential for developing a blood-related disorder, comprising the steps of:
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(a) isolating genomic DNA from a human individual; (b) assaying said DNA for a deletion relative to DNA of a normal human subject, said deletion reducing the number of α
-globin genes in the genome of the human individual; and(c) determining an increased potential for developing a blood-related disorder in said human individual, wherein the presence of said deletion in the genomic DNA of the human individual is indicative of an increased potential for developing said blood-related disorder, said blood-related disorder selected from the group consisting of hypertension, myocardial infarction, and diabetes. - View Dependent Claims (24)
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25. A kit comprising:
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an assay means for assaying genomic DNA from a human subject for the presence of an α
-globin deletion mutation; anda means for correlating the presence of an α
-globin deletion mutation to an increased potential of developing a blood-related disorder, said disorder selected from the group consisting of hypertension, myocardial infarction, and diabetes. - View Dependent Claims (26, 27, 28)
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29. An assay for identifying a mild α
- -thalassemia deletion genotype in a human individual comprising the steps of;
(a) providing genomic DNA from said human individual; (b) performing a multiplex polymerase chain reaction with said genomic DNA to amplify a marker portion and a control portion thereof, wherein the presence of an α
-globin gene deletion in said genomic DNA reduces the quantity of marker amplicons produced in the multiplex polymerase chain reaction relative to the quantity of control amplicons produced in said multiplex polymerase chain reaction;(c) determining a ratio of marker amplicons to control amplicons produced in the multiplex polymerase chain reaction; (d) comparing the ratio of step (c) to a ratio of marker amplicons to control amplicons produced in a multiplex polymerase chain reaction performed with genomic DNA of a normal human subject; and (e) identifying a mild α
-thalassemia deletion genotype in said human individual from said ratio of step (c), wherein a mild α
-thalassemia genotype in said individual correlates with a lower marker amplicon to control amplicon ratio of step (c) compared to the ratio of marker amplicons to control amplicons produced in a multiplex polymerase chain reaction performed with genomic DNA of a normal human subject. - View Dependent Claims (30, 31, 32, 33)
- -thalassemia deletion genotype in a human individual comprising the steps of;
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34. A method of determining a hypertension-correlated genetic disorder in a hypertensive human individual comprising the steps of:
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(a) isolating DNA from said human individual; (b) assaying said DNA for a deletion relative to DNA of a normal human subject, said deletion comprising a reduction in the number of α
-globin genes in the genome of the hypertensive human individual; and(c) determining a presence of a hypertension-correlated genetic disorder in said individual from a presence of said deletion in said DNA.
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35. A kit for determining the α
- -globin genotype of a human individual comprising, in association;
a first pair of oligonucleotide primers for amplifying a first marker portion of a human chromosome 16, said first marker portion being absent from a human chromosome 16 having a single α
-gene deletion selected from the group consisting of an -α
3.7 deletion and an -α
4.2 deletion;a second pair of oligonucleotide primers for amplifying a second marker portion of a human chromosome 16, said second marker portion being present in a human chromosome 16 having the single α
-gene deletion and being absent from a human chromosome 16 having an --MED deletion; anda control pair of oligonucleotide primers for amplifying a control portion of human genomic DNA, wherein said first pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQ ID NO;
4 and a primer having the nucleotide sequence of SEQ ID NO;
5; and
wherein said second pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQ ID NO;
6 and a primer having the nucleotide sequence of SEQ ID NO;
7.
- -globin genotype of a human individual comprising, in association;
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36. A kit for determining the α
- -globin genotype of a human individual comprising, in association;
a first pair of oligonucleotide primers for amplifying a first marker portion of a human chromosome 16, said first marker portion being absent from a human chromosome 16 having an -α
3.7 deletion;a second pair of oligonucleotide primers for amplifying a second marker portion of a human chromosome 16, said second marker portion being present in a human chromosome 16 having the single α
-gene deletion and being absent from a human chromosome 16 having an --MED deletion;a third pair of oligonucleotide primers for amplifying a third marker portion of a human chromosome 16, said third marker portion being absent from a human chromosome 16 having an -α
4.2 deletion; anda control pair of oligonucleotide primers for amplifying a control portion of human genomic DNA, wherein said first pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQ ID NO;
4 and a primer having the nucleotide sequence of SEQ ID NO;
5;
wherein said second pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQ ID NO;
6 and a primer having the nucleotide sequence of SEQ ID NO;
7; and
wherein said third pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQ ID NO;
12 and a primer having the nucleotide sequence of SEQ ID NO;
13.
- -globin genotype of a human individual comprising, in association;
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37. A kit for determining the α
- -globin genotype of a human individual by multiplex polymerase chain reaction with genomic DNA of said individual, said kit comprising, in association;
a first pair of oligonucleotide primers for amplifying a first marker portion of a human chromosome 16, said first marker portion being absent from a human chromosome 16 having an -α
3.7 deletion wherein said first pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQUENCE ID NO;
4 and a primer having the nucleotide sequence of SEQUENCE ID NO;
5;a second pair of oligonucleotide primers for amplifying a second marker portion of a human chromosome 16, said second marker portion being present in a human chromosome 16 having a single α
-gene deletion selected from the group consisting of an -α
3.7 deletion and an -α
4.2 deletion and being absent from a human chromosome 16 having an --MED deletion;a third pair of oligonucleotide primers for amplifying a third marker portion of a human chromosome 16, said third marker portion being absent from a human chromosome 16 having an -α
4.2 deletion wherein said third pair of oligonucleotide primers consists of a primer having the nucleotide sequence of SEQUENCE ID NO;
12 and a primer having the nucleotide sequence of SEQUENCE ID NO;
13; anda control pair of oligonucleotide primers for amplifying a control portion of human genomic DNA, said primer pairs permitting a determination, via multiplex polymerase chain reaction, of a -α
3.7 α
/α
α
mild α
-thalassemia genotype, a -α
4.2 α
/α
α
mild α
-thalassemia genotype, and α
-thalassemia trait genotypes wherein at least two α
-globin gene alleles have been deleted. - View Dependent Claims (38, 39)
- -globin genotype of a human individual by multiplex polymerase chain reaction with genomic DNA of said individual, said kit comprising, in association;
Specification