WO2021226581A1 - Multidimensional barcode pooling - Google Patents

Abstract

The invention provides a method for multiplexed screening of subject samples for two or more target conditions. The method may include dividing a set of samples comprising nucleic acid markers for at least two target conditions for analysis into at least two sets of pools comprising a first set of X pools and a different second set of Y pools, wherein a subsample from every sample in the set of samples is included in the at least two sets of pools, wherein: X is the number of pools into which the samples are divided for the first set of pools; and Y is the number of pools into which the samples are divided for the second set of pools; and X and Y establish a pool matrix in which every sample in the set of samples has an intersection between the X pools and the Y pools. The method may include coupling a nucleic acid barcode to the nucleic acid markers of each pool to yield barcoded nucleic acid markers, wherein the barcode is unique to the pool and differentiated from all other pools in the at least two sets of pools.

Description

MULTIDIMENSIONAL BARCODE POOLING

Related Applications

[00001] This application claims priority to and the benefit of U.S. Provisional Patent App. No. 63/021,985 filed on May 8, 2020 and U.S. Provisional Patent App. No. 63/161,747 filed on March 16, 2021; the disclosures of which are hereby incorporated herein by reference.

Field of the Invention

[00002] The invention relates to the field of screening subject samples for rare and higher frequency diseases or conditions.

Background of the Invention

[00003] There are an increasing number of diseases for which individuals, such as newborns, can be screened. For example, there are about 7,000 rare genetic diseases afflicting millions of people which have thus far been described. A number of these diseases strike very early in life putting a heavy burden on the pediatric population and in particular, the newborn population.

[00004] To date, efforts for screening newborns for genetic diseases have focused on a relatively small number of conditions (e.g., about 50 or less conditions). In contrast, efforts by pharmaceutical companies to develop molecular agents to treat disease have led to an increasing number of genetic conditions that can be described as treatable. This disparity has created a gap between newborn screening practices and the number of available therapies.

[00005] Diagnosis of a rare disease can be a lengthy process which typically involves documenting clinical manifestations and performing imaging and/or biochemical tests. For newborns, certain conditions demand immediate diagnosis and initiation of treatment within hours of birth before devastating and often irreversible health consequences occur. The relatively recent application of next generation sequencing (NGS) technologies to the diagnostic workflow for a rare disease has provided a powerful alternative for genetic analysis in the early stages of the disease process. Flowever, many clinical gene sequencing providers have turnaround times of about 3 weeks or more, and the per-sample cost for sequencing is relatively high. The long turnaround time and high sequencing costs can be a barrier to using NGS technologies in large screening population applications like newborn screening. [00006] There is a need in the art for ways to reduce the costs associated with sequencing and screening markers of disease. There is a need in the art for ways to reduce the time from sample collection to presentation of results associated with sequencing and screening markers of disease.

Summary of the Invention

[00007] The invention provides a method for multiplexed screening of subject samples for two or more target conditions. The method may include dividing a set of samples including nucleic acid markers for at least two target conditions for analysis into at least two sets of pools. The pools may include a first set of X pools and a different second set of Y pools, wherein a subsample from every sample in the set of samples may be included in the at least two sets of pools, wherein X is the number of pools into which the samples may be divided for the first set of pools; and Y is the number of pools into which the samples may be divided for the second set of pools; and X and Y establish a pool matrix in which every sample in the set of samples has an intersection between the X pools and the Y pools. The method may include coupling a nucleic acid barcode to the nucleic acid markers of each pool to yield barcoded nucleic acid markers, wherein the barcode is unique to the pool and differentiated from all other pools in the at least two sets of pools. The method may include sequencing the nucleic acid markers from the sets of pools, wherein each sample may be sequenced at least once in the X pools and at least once in the Y pools. The method may include using a computer, deconvoluting the sequencing data by identifying the markers at the intersections of the pool matrix for which the sequencing data may be indicative of the target condition.

[00008] Sequencing the nucleic acid markers from the sets of pools may include pooling the sets of pools into a set of one or more master pools and may include sequencing the nucleic acid markers from the set of master pools to produce the sequencing data. In some cases, pooling the sets of pools into the set of one or more master pools produces one master pool. In some cases, pooling the sets of pools into the set of one or more master pools produces two master pools. In some cases, pooling the two sets of pools into the set of one or more master pools produces three master pools.

[00009] In some cases, X greater than 12. In some cases, X and Y may each be greater than 12. In some cases, X is greater than 20. In some cases, X is greater than 20 and Y is greater than 12. In some cases, X and Y may each be greater than 20. In some cases, X is greater than 50 and Y is greater than 12. In some cases, X and Y may each be greater than 50.

[00010] In certain embodiments, the set of samples originate from a child. In certain embodiments, the set of samples originate from an infant. In certain embodiments, the set of samples originate from a newborn infant. In certain embodiments, the sample includes nucleic acid markers obtained from blood, plasma, serum, urine, or another bodily fluid. In certain embodiments, the sample includes nucleic acid markers obtained from a reconstituted blood spot.

[00011] In certain embodiments, the set of samples include nucleic acid markers for at least ten target conditions. In certain embodiments, the set of samples include nucleic acid markers for at least 50 target conditions. In certain embodiments, the set of samples include nucleic acid markers for at least 100 target conditions. In some cases the target conditions include the conditions set forth in Table 1. In some cases the target conditions include conditions selected from Table 2. In some cases the target conditions include conditions selected from Table 3. In some cases the target conditions include at least 10 conditions selected from Table 2. In some cases the target conditions include at least 10 conditions selected from Table 3. In some cases the target conditions include at least 20 conditions selected from Table 2. In some cases the target conditions include at least 20 conditions selected from Table 3. The target conditions may be selected from combinations of the foregoing tables.

[00012] In certain embodiments, the sequencing data may be referenced against a reference database of mutations and associated probabilities of conditions to identify a probability that the sequencing data may be indicative of the target condition. In certain embodiments, the database may be maintained as part of a system in which the identification of the markers indicative of the target condition may be correlated with subsequent empirical data from a sample and a probability of the target condition based on the empirical data may be updated in the database.

[00013] In certain embodiments, a computer may be used to provide an output indicating when the target condition may not be distinguishable between positive and carrier status. In certain embodiments, the computer output may include a recommendation for subsequent testing to make a definitive determination of positive or carrier status. The method may include carrying out the subsequent testing, wherein subsequent testing includes sequencing a nucleic acid sample from a subject.

[00014] The method may further include using the computer to provide an output indicating when the deconvoluting of the sequencing data does not clearly identify individual samples from a master pool for which the sequencing data may be indicative of the target condition. In some cases, the computer may recommend subsequent testing to make a definitive determination of an individual sample for which the sequencing data may be indicative of the target condition. The method may further include carrying out the subsequent testing, wherein the subsequent testing may include sequencing the nucleic acid sample from one or more subjects from the master pool from which the sequencing data does not clearly identify individual samples. [00015] In certain embodiments, the nucleic acid markers for analysis and the at least two target conditions may be selected to result in a successful deconvolution rate of at least 50%. In certain embodiments, the nucleic acid markers for analysis and the at least two target conditions may be selected to result in a successful deconvolution rate of at least 70%. In certain embodiments, the nucleic acid markers for analysis and the at least two target conditions may be selected to result in a successful deconvolution rate of at least 95%.

[00016] In certain embodiments, the samples may be from a testing population and the target conditions may include rare conditions which may have an incidence rate in the testing population of less than 1 in 50. In certain embodiments, the samples may be from a testing population and the target conditions may include rare conditions which may have an incidence rate in the testing population of less than 1 in 200. In certain embodiments, the samples may be from a testing population and the target conditions may include rare conditions which may have an incidence rate in the testing population of less than 1 in 500.

[00017] In certain embodiments, the samples may be from a testing population and at least 50% of the target conditions may have an incidence rate in the testing population of less than 1 in 50. In certain embodiments, the samples may be from a testing population and at least 75% of the target conditions may have an incidence rate in the testing population of less than 1 in 200. In certain embodiments, the samples may be from a testing population and at least 90% of the target conditions may have an incidence rate in the testing population of less than 1 in 500.

[00018] In certain embodiments, the samples may be from a testing population and X and Y may each be at least 10 and at least 50% of the target conditions may have the incidence rate in the testing population of less than 1 in 50. In certain embodiments, the samples may be from a testing population and X and Y may each be at least 50 and at least 75% of the target conditions may have the incidence rate in the testing population of less than 1 in 200. In certain embodiments, the samples may be from a testing population and X and Y may each be at least 200 and at least 90% of the target conditions may have the incidence rate in the testing population of less than 1 in 500.

[00019] In some cases, the target conditions may include at least 10 conditions. In some cases X and Y may each be greater than 10. In some cases at least 90% of the target conditions may have an incidence rate in the testing population of less than 1 in 50. In some embodiments, the target conditions may include at least 50 conditions. In some embodiments, the sets of pools may include samples from at least 50 subjects. In some embodiments, at least 90% of the target conditions may have an incidence rate in the testing population of less than 1 in 200. In some cases the target conditions may include at least 200 conditions. In some cases, X and Y may each be greater than 200. In some cases at least 90% of the target conditions may have an incidence rate in the testing population of less than 1 in 500.

[00020] In certain embodiments, some or all of the markers may be selected to give a yes/no answer with respect to one or more of the target conditions. In certain embodiments, some or all of the markers may be selected to give a probability of a yes or a probability of a no with respect to one or more of the target conditions.

[00021] In certain embodiments, the target conditions may include a disease for which newborns may be screened. In certain embodiments, the target conditions may all be diseases for which newborns may be screened. In certain embodiments, the target conditions may include an enzyme deficiency or an enzyme activity deficiency. In certain embodiments, the target conditions may include an enzyme deficiency and an enzyme activity deficiency. In certain embodiments, the target conditions may include a metabolic disorder. In certain embodiments, the target conditions may include an enzyme deficiency and an enzyme activity deficiency and a metabolic disorder.

Brief Description of the Drawings

[00022] FIG. 1 illustrates a flow diagram of an example method of using a one-dimensional (ID) pooling strategy for multiplexed sample screening.

[00023] FIG. 2 shows pictorially one of the steps of the method of FIG. 1.

[00024] FIG. 3 illustrates a flow diagram of an example method of using a two-dimensional (2D) pooling strategy for multiplexed sample screening.

[00025] FIG. 4A and 4B show pictorially two steps of the method of FIG. 3.

[00026] FIG. 5 is a table showing another example of a two-dimensional pooling strategy.

[00027] FIG. 6 illustrates a schematic diagram of positive hits in a two-dimensional pooling strategy for an autosomal recessive variant.

[00028] FIG. 7 illustrates a flow diagram of an example workflow for sequencing and screening markers of disease.

[00029] FIG. 8 illustrates a flow diagram of an example of a method of extracting genomic DNA from a reconstituted dried blood spot.

[00030] FIG. 9 is a table showing the yield metrics for DNA extracted from dried blood spot punches using the method of FIG. 8. [00031] FIG. 10 illustrates a flow diagram of an example of a method of preparing a sequencing library from pooled genomic DNA.

[00032] FIG. 11 illustrates how the empirical and theoretical estimates for carrier frequency are obtained.

[00033] FIG. 12 is a plot showing positive "hits" identified in a two-dimensional pooling strategy for a low frequency variant (VI) and a high frequency variant (V2).

[00034] FIG. 13 is a table showing the computer simulated two-dimensional screening of 100,000 newborns based on GAA variant frequency.

[00035] FIG. 14A is a plot of the frequency of incorrect hits per run for the rare variant panel of FIG. 13.

[00036] FIG. 14B is a plot of the common variant panel of FIG. 13.

[00037] FIG. 15 illustrates a schematic diagram of the expected and experimentally observed hits for a high frequency variant GAA c.-32-13T>G in a two-dimensional screening.

[00038] FIG. 16A and FIG. 16B illustrate a one-dimensional screening strategy for detecting known GAA variants in 2 sets of pooled samples.

[00039] FIG. 17A is a plot of expected variant coverage in the first set of n = 10 unpooled samples of FIG. 16A versus observed variant coverage in the pooled samples.

[00040] FIG. 17B is a plot of expected variant coverage in the first set of n = 20 unpooled samples of FIG 16A versus observed variant coverage in the pooled samples.

[00041] FIG. 18A is a plot of expected variant coverage in the second set of n = 10 unpooled samples of FIG. 16B versus observed variant coverage in the pooled samples.

[00042] FIG. 18B is a plot of expected variant coverage in the second set of n = 20 unpooled samples of FIG. 16B versus observed variant coverage in the pooled samples.

[00043] FIG. 19A is a plot of a comparison of variant coverage within the n = 10 DNA pooled experimental replicates of FIG. 16A.

[00044] FIG. 19B is a plot of a comparison of variant coverage within the n = 20 DNA pooled experimental replicates of FIG. 16A.

[00045] FIG. 19C is a plot of a comparison of variant dropouts between the n = 10 and n = 20 DNA pooled experimental replicates of FIG. 16A. Abbreviations and Definitions

[00046] "ID" is the abbreviation for one dimensional.

[00047] "2D" is the abbreviation for two-dimensional.

[00048] "CNV" is the abbreviation for copy number variants.

[00049] "LP" is the abbreviation for likely pathogenic variant.

[00050] "P" is the abbreviation for pathogenic variant.

[00051] "SNV" is the abbreviation for single nucleotide variant.

[00052] "VI" is the abbreviation for low frequency variant.

[00053] "V2" is the abbreviation for high frequency variant.

[00054] "Aligned reads" means the percentage of all sequence reads generated for a sample that align to the reference genome.

[00055] "Decoding accuracy of variant calls" means concordance or decoding accuracy of variant calls.

[00056] "Enrichment" means the percentage of all sequence reads generated for a sample that align to the target regions for the assay.

[00057] "Indicative of a target condition" means that the target condition is present in a subject sample for analysis or that the results of the method of the invention indicate the possibility of the target condition being present in a subject sample. For example, a finding relating to a target condition that does not distinguish between a carrier or an actual positive would be indicative of the target condition. Similarly, a finding in which a positive finding is produced in one dimension of a matrix but is absent in another dimension of a matrix the intersection of pools would be indicative of the target condition. Individuals whose samples are identified as indicative of a target condition may be referred for subsequent testing. The methods of the invention may include conducting such subsequent testing.

[00058] "Intersecting position" means a point in the intersection of pools at which barcodes from intersecting samples are positive.

[00059] "Markers" of a disease, condition or disorder include, but are not limited to, the captured probes target regions provided in Table 2 and Table 3.

[00060] "Matrix" means a theoretical array of data. The array may be for example, a ID array laid out in one-dimensional space with only an X axis. The array may be for example, a 2D X, Y array, laid out in two-dimensional space with an X axis and a Y axis. The array may be for example, a 3D X, Y, Z array, laid out in three-dimensional space with an X axis and a Y axis. The array may be multidimensional. An "array" is theoretical in the sense that the data must be structured or provided in a manner that would permit an actual array to be computed from the data, but it is not necessary for the array to actually be created. Thus, for example, a 2D or 3D array may be provided using data that is not arranged in an actual array (e.g., a list of array positions, or a list of array positions associated with a list of values, such as test values or barcodes), so long as the theoretical array could be computed from the data.

[00061] "Mean coverage" means the mean coverage for the pooled sample across the target region.

[00062] "Pool" means a combined set of nucleic acid markers from multiple subject samples. In some cases, the combined set of nucleic acid markers is from multiple subjects.

[00063] "Sample" means a sample comprising a nucleic acid, such as RNA or DNA. The nucleic acid may be enriched for specific targets using various known techniques, such as pull-down by probes or amplification by targets.

[00064] "Sample source" means the origin of the nucleic acid in the sample. For example, the sample source is blood, plasma, serum, urine, or another bodily fluid. In one aspect, the sample is a reconstituted blood spot.

[00065] "Subject" means the individual being tested.

[00066] "Testing population" means the subset of the population from which the test subjects are selected, e.g., the population of newborn infants would be a test population for newborn screening tests.

Detailed Description of the Invention

[00067] The invention provides a system and methods for multiplexed screening of subject samples to identify individual samples that are positive or negative for specific markers or combinations of markers of a disease, condition or disorder.

[00068] The invention provides a method for multiplexed screening of subject samples for two or more target conditions. The method may include dividing a set of samples comprising nucleic acid markers for at least two target conditions for analysis into at least two sets of pools comprising a first set of X pools and a different second set of Y pools, wherein a subsample from every sample in the set of samples is included in the at least two sets of pools, wherein: X is the number of pools into which the samples are divided for the first set of pools; and Y is the number of pools into which the samples are divided for the second set of pools; and X and Y establish a pool matrix in which every sample in the set of samples has an intersection between the X pools and the Y pools. The method may include coupling a nucleic acid barcode to the nucleic acid markers of each pool to yield barcoded nucleic acid markers, wherein the barcode is unique to the pool and differentiated from all other pools in the at least two sets of pools. The method may include sequencing the nucleic acid markers from the sets of pools, wherein each sample is sequenced at least once in the X pools and at least once in the Y pools. The method may include using a computer, deconvoluting the sequencing data by identifying the markers at the intersections of the pool matrix for which the sequencing data is indicative of the target condition.

One-dimensional (ID) Pooling Strategy

[00069] In one aspect, a method of the invention uses a one-dimensional (ID) pooling strategy, wherein DNA extracts from multiple subject samples (e.g., 40 subject samples) are combined in two or more pools of samples (e.g., 4 pools each containing 10 subject samples) with molecular barcodes and used to generate x-number of pooled-barcoded libraries. The x-number of pooled-barcoded libraries are then combined into a single sample and sequenced. Pools with a positive result or "hit" for a specific marker or combinations of markers are identified and the individual subject samples in the positive pool are sequenced to identify the positive subject sample. A positive result or hit means that one or more samples in a pool contain a pathogenic variant (P) or a likely pathogenic (LP) variant.

[00070] FIG. 1 illustrates a flow diagram of an example of a method 100 of using a one-dimensional (ID) pooling strategy for multiplexed sample screening. In this example, multiple samples (e.g., 40 different subject samples) are sequenced in a single run, and pools with positive results or hits are then sequenced individually. Method 100 may include, but is not limited to, the following steps:

[00071] At a step 110, DNA extracts from multiple subject samples are obtained. In one example, DNA extracts from 40 different subjects are obtained.

[00072] At a step 115, the DNA samples are combined into pools and individual pooled-barcoded libraries are prepared using unique barcodes for each pool. For example, the 40 different DNA samples are combined into 4 pools each containing 10 DNA samples (n = 10). A pooled-barcoded sequencing library is then prepared from each pool using unique barcodes. In this example, 4 unique barcodes are used.

[00073] At a step 120, the individual pooled-barcoded libraries are combined into one sample and sequenced. In one example, the pooled-barcoded libraries (i.e., 4 barcoded libraries of 10 DNA samples each) are combined into a single sample (i.e., n = 40 subject samples) and sequenced using lllumina's MiniSeq technology, for example.

[00074] At a step 125, pools with positive "hits" are identified. For example, the sequencing data from the combined pooled-barcoded libraries is deconvoluted using the unique pool-specific barcodes to provide sequencing data at the per pool level.

[00075] At a step 130, individual subject samples represented in a positive pool are sequenced to determine which subject sample in the pool contains the pathogenic variant(s) or likely pathogenic variant(s).

[00076] FIG. 2 shows pictorially one of the steps of method 100 of FIG. 1. Namely, in step 115, the 40 different DNA samples are divided into 4 pools, i.e., pools 210, 215, 220, and 225, wherein each pool includes 10 different subject samples (n = 10). A pooled-barcoded library is then prepared from each pool using unique barcodes (indicated by the different fill colors).

Two-Dimensional (2D) Pooling Strategy

[00077] In one aspect, a method of the invention uses a two-dimensional (2D) pooling strategy of x- number of rows and y-number of columns, wherein each subject sample is labeled with two separate pool-level barcodes, and positive results of individual subject samples are identified at the "intersection" of the pools.

[00078] FIG. 3 illustrates a flow diagram of an example method 300 of using a two-dimensional (2D) pooling strategy for multiplexed sample screening. Method 300 may include, but is not limited to, the following steps:

[00079] At a step 310, DNA extracts from multiple subject samples are obtained. In one example, DNA extracts from 40 different subjects are obtained.

[00080] At a step 315, the DNA samples are divided into 2 sets of pools and z-number of pooled- barcoded libraries are prepared using unique barcodes for each pool. For example, the 40 different DNA samples are divided into a first set of 4 pools where each pool includes 10 DNA samples and a second set of 10 pools where each pool includes 4 DNA samples. A pooled-barcoded sequencing library is then prepared from each pool using pool-specific unique barcodes. In this example, 14 unique barcodes are used to prepare z = 14 libraries.

[00081] At a step 320, the pooled-barcoded libraries are combined into one sample and sequenced. In one example, the pooled-barcoded libraries (i.e., 14 barcoded libraries representing 40 subject DNA samples) are combined into one sample and sequenced using lllumina's MiniSeq technology, for example. [00082] At a step 325, subject samples with positives are identified. For example, the sequencing data from the combined pooled-barcoded libraries is deconvoluted using the pool-specific barcodes to provide sequencing data at the per pool level. Subject samples in a positive pool are then identified using the unique combination of barcodes. A positive means the sample contains the pathogenic variant(s) or likely pathogenic variant(s).

[00083] FIG. 4A and 4B show pictorially two of the steps of method 300 of FIG. 3. With reference to FIG. 4A, in step 315 the 40 different DNA samples are combined into 2 sets of pools, a first set in the row dimension of a matrix (labeled A, B, C, and D) and the second set in the column dimension of the matrix (labeled "a" through "j"). Each pool in the first set of pools (i.e., A through D) includes 10 DNA samples. Each pool in the second set of pools (i.e., "a" through "j") includes 4 DNA samples. A pooled-barcoded sequencing library is prepared from each pool using unique pool-specific barcodes. In this example, 14 unique pool-specific barcodes are used to prepare 14 libraries (i.e., 4 barcoded libraries in the row dimension and 10 barcoded libraries in the column dimension).

[00084] With reference to FIG. 4B, in step 325 the sequencing data from the combined pooled- barcoded libraries is deconvoluted using the pool-specific barcodes to identify positive pools (indicated by arrows at pool "B" and pool "d"). The individual positive subject sample is then identified by the unique combination of barcodes at the intersection of row level barcode and column level barcode. In this example a positive subject sample (indicated by circle) is identified by the barcode for pool "B" and the barcode for pool "d", i.e., B/d. A positive means the sample contains the pathogenic variant(s) or likely pathogenic variant(s).

[00085] FIG. 5 is a table 500 showing another example of a two-dimensional pooling strategy. In this example, the top row identifies a first set of pools and the left column identifies a second set of pools, and each of the remaining cells identifies a unique first/second pool level barcode, A/B. For example, in this example, 25 samples can be sequenced in a single sequencing run and individual positives can be identified at the intersections of the pools. For example, a positive in pool A3 and B2 indicates a positive in the unique sample 3/2, highlighted in table 500.

[00086] FIG. 6 illustrates a schematic diagram of positives in a two-dimensional (2D) pooling strategy for an autosomal recessive variant. For an autosomal recessive variant, carrier frequencies are greater than the frequency for an affected case.

[00087] The deconvolution rate is the proportion of samples for which carrier or affected status can be resolved from the first level 2D pooled testing. In some cases, markers and the at least two target conditions selected for a method of the invention, are selected to affect a deconvolution rate of at least 50, 60, 70, 80, 90, or 95%. Pooling Strategy Parameters

[00088] Parameters that can be varied in a pooling strategy (e.g., a two-dimensional pooling strategy) include, for example, the number of conditions that can be targeted for screening, selection of variants based on their incidence rate in the population (e.g., variants in a rare condition), and number and size of pools.

[00089] In one aspect, the conditions selected include conditions characterized by single nucleotide variants.

[00090] In one aspect, the conditions selected include copy number variants.

[00091] In one aspect, the conditions selected include both single nucleotide variants and copy number variants, where copy number variants include insertions, multiplications, deletions, and/or translocations.

[00092] In one aspect, the method includes testing for 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400 or more target conditions, where each target condition may include testing for one or more target markers.

[00093] In one aspect, the target conditions are rare conditions, such as conditions having an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00094] In one aspect, at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00095] In one aspect, at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00096] In one aspect, at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00097] In one aspect, the test pools of at least 10 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500. [00098] In one aspect, the test pools of at least 10 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00099] In one aspect, the test pools of at least 10 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00100] In one aspect, the test pools of at least 25 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00101] In one aspect, the test pools of at least 25 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00102] In one aspect, the test pools of at least 25 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00103] In one aspect, the test pools of at least 40 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00104] In one aspect, the test pools of at least 40 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00105] In one aspect, the test pools of at least 40 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00106] In one aspect, the test pools of at least 50 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00107] In one aspect, the test pools of at least 50 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500. [00108] In one aspect, the test pools of at least 50 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00109] In one aspect, the test pools of at least 75 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00110] In one aspect, the test pools of at least 75 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00111] In one aspect, the test pools of at least 75 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00112] In one aspect, the test pools of at least 100 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00113] In one aspect, the test pools of at least 100 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00114] In one aspect, the test pools of at least 100 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00115] In one aspect, the test pools of at least 200 subjects and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00116] In one aspect, the test pools of at least 200 subjects and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00117] In one aspect, the test pools of at least 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00118] In one aspect, the test includes at least 10 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00119] In one aspect, the test includes at least 25 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00120] In one aspect, the test includes at least 50 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00121] In one aspect, the test includes at least 75 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00122] In one aspect, the test includes at least 100 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00123] In one aspect, the test includes at least 200 conditions, pools at least 10, 40, 50, 100, or 200 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500.

[00124] In one aspect, the test includes at least 10 conditions, pools at least 10 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50.

[00125] In one aspect, the test includes at least 25 conditions, pools at least 25 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 100.

[00126] In one aspect, the test includes at least 25 conditions, pools at least 40 subjects and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 150.

[00127] In one aspect, the method includes testing for 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400 or more target markers.

[00128] In one aspect, the method includes testing for 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150,

200 or more samples in a single run. [00129] In one aspect, the samples comprise nucleic acid markers of 10 or more target conditions. In another aspect, the samples collectively comprise nucleic acid markers of 10 or more target conditions.

[00130] In one aspect, the samples comprise nucleic acid markers of 50 or more target conditions. In another aspect, the samples collectively comprise nucleic acid markers of 50 or more target conditions.

[00131] In one aspect, the samples comprise nucleic acid markers of 100 or more target conditions.

In another aspect, the samples collectively comprise nucleic acid markers of 100 or more target conditions.

[00132] In some aspects, some or all of the markers are selected to give a yes/no answer with respect to a target condition. In some aspects, some or all of the markers are selected to give a probability with respect to a target condition. In some aspects, some or all of the markers are selected to give at least an X probability with respect to a target condition, where X = 50%, 60%,

70%, 80%, 90%, 95%, 99%, 99.9%. In some aspects, at least 50% of the markers are selected to give at least an X probability with respect to a target condition, where X = 50%, 60%, 70%, 80%, 90%,

95%, 99%, 99.9%. In some aspects, at least 75% of the markers are selected to give at least an X probability with respect to a target condition, where X = 50%, 60%, 70%, 80%, 90%, 95%, 99%, or 99.9%. In some aspects, at least 90% of the markers are selected to give at least an X probability with respect to a target condition, where X = 50%, 60%, 70%, 80%, 90%, 95%, 99%, or 99.9%. In some aspects, at least 90% of the markers are selected to give at least an X probability with respect to a target condition, where X = 90%, 95%, 99%, or 99.9%.

Conditions

[00133] In one aspect, the condition is a disease for which newborns are screened. In one aspect, the condition is an enzyme deficiency or an enzyme activity deficiency. In one aspect, the condition is a metabolic disorder. In one aspect, the condition is a metabolic disorder having an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500. In one aspect, the condition is a metabolic disorder, and the panel includes at least 50%, at least 75%, or at least 90% of conditions having an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500. In one aspect, the condition is a metabolic disorder, and the panel includes at least 10, 20, 30, 40, 50, 100, 200, 300, or 400 conditions having an incidence rate in the testing population of less than 1 in 50, 1 in 100, or less than 1 in 200, or less than 1 in 300, or less than 1 in 400, or less than 1 in 500. [00134] In one aspect, the condition is selected from the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 25 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi dimensional pooled samples include tests for at least 75 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 100 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 150 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 175 of the disorders identified in the list in Table 1.

[00135] In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 150 disorders including tests for at least 100 of the disorders identified in the list in Table 1. In one aspect, the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1.

[00136] In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 20 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1.

[00137] In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 40 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1. [00138] In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 60 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1.

[00139] In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 80 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1.

[00140] In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 20 disorders including tests for at least 10 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 40 disorders including tests for at least 20 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 60 disorders including tests for at least 40 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 80 disorders including tests for at least 60 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 80 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 100 disorders including tests for at least 50 of the disorders identified in the list in Table 1. In one aspect, the multi-dimensional pooled samples include at least 100 pooled samples and the tests conducted on the multi-dimensional pooled samples include tests for at least 200 disorders including tests for at least 100 of the disorders identified in the list in Table 1.

[00141] In the aspects described herein, generally speaking, the method involves sample preparation to add the barcode combinations, pooling, sequencing, and deconvoluting to identify individual samples that are positive or negative for specific markers or combinations of markers.

[00142] Although the method is described with reference to one-dimensional or two-dimensional pooling, it may be appreciated that more dimensions can be used, e.g., three-dimensional pooling using three barcodes per sample, four-dimensional pooling using four barcodes per sample, etc. The number of dimensions used in a pooling strategy may be selected based on the frequency of variants in a population. For example, a three-dimensional pooling strategy may be used for conditions that have variant frequencies between X and Y (e.g., common variants in a population); and a two- dimensional pooling strategy may be used for conditions that have variant frequencies less than X (e.g., rare variants in a population).

[00143] A pooling strategy may be selected based on variant frequencies and used to reduce the number of incorrect hit rates (overlapping, unresolvable hits) such that resolution of a correct positive(s) is at the sample level.

[00144] A pooling strategy may be selected based on variant frequencies and the number of conditions to be screened.

[00145] The invention makes use of in silico deconvolution in which each marker or set of markers for a target condition is analyzed across multiple pool matrices, e.g., one or more matrices per condition.

[00146] In some cases, individuals with positive results are referred for further individual testing.

[00147] In some cases, samples from individuals with positive results are individually tested to confirm the results.

[00148] The invention includes a system including a reference database of mutations and associated probabilities of conditions. In operation, the database is referenced based on results of sequence analysis to identify probabilities of disease based on specific mutation profiles identified in the analysis.

[00149] The system of the invention may be a learning system, in which data from subsequent testing or other diagnostic analyses is fed back into the system to improve subsequent results.

[00150] In some aspects, one or more of the conditions tested for may not permit distinguishing of positive (affected) versus carrier status, and subsequent testing may be recommended or carried out to make a more definitive determination. Second level testing to resolve affected versus carrier status can be done, for example, through targeted next-generation sequencing or PCR.

Methods

[00151] The following example illustrates an example of the methods of the invention. It will be appreciated that variations and alternatives will be acceptable within the scope of the invention.

[00152] FIG. 7 illustrates a flow diagram example of a workflow 700 for sequencing and screening markers of disease. Workflow 700 includes, but is not limited to, the following steps:

[00153] In a step 710, a pooled-barcoded sequencing library is prepared and sequenced. For example, a sequencing library is prepared using genomic DNA extracted from reconstituted dried blood spots and sequenced as a paired end (2x75bp) run on the lllumina MiniSeq Sequencing by Synthesis (SBS) platform.

[00154] In a step 715, the sequencing data is aligned, and variants are called.

[00155] In a step 720, sequencing quality control (QC) procedures to evaluate the sequencing data are performed. For example, QC protocols that can be performed include, but are not limited to, evaluation of raw sequencing data quality and depth, alignment quality, and reads duplication rates.

[00156] In a step 725, variant filtering is performed. For example, protocols are followed to filter out the incorrectly called variants while retaining the true variant calls.

[00157] In a step 730, reports are generated. For example, for each subject sample in the sequence library, a preliminary report of a positive (e.g., pathogenic (P) or likely pathogenic (LP)) or negative hit for each marker tested is generated. For positive variant calls, a request for second tier screening and/or a clinical consultation report is generated.

[00158] In one example, a sequencing library is prepared using genomic DNA extracted from reconstituted dried blood spots. The extraction of genomic DNA from a reconstituted dried blood spot is, for example, performed in a deep-well microtiter plate, where the shape of the wells and profile of well bottoms are selected to provide high recovery of genomic DNA from the dried blood spot sample. In one example, the wells of the plate have a square shape and well bottoms have a pyramid or V-shape.

[00159] Existing protocols and/or kits for extracting genomic DNA from dried blood spot punches typically use 1 to 3 dried blood spot punches per extraction and may include a denaturation step (e.g., 90°C or higher) and an overnight incubation period. For example, a DBS extraction kit available from Norgen Biotedk typically uses three - 3 mm punches and yields an average of about 150 ng of DNA; a DBS extraction kit available from Geneaid typically uses one - 6 mm punch and yields an average of about 300 ng of DNA; a DBS extraction kit available from Qiagen typically uses three - 3 mm punches and yields an average of about 75 ng of DNA; and a DBS extraction kit available from GenTegra typically uses one - 6 mm punch and yields an average of about 50 ng of DNA.

[00160] The invention makes use of a method for extracting genomic DNA from 1, 2, or 3 dried blood spot punches in about 2 hours. In one example, a dried blood spot punch has a diameter of 3.2 mm. In a preferred embodiment, a single 3.2 mm diameter dried blood spot punch is used for extraction of genomic DNA. Flowever, it will be appreciated that two or more punches can be combined for a single extraction. [00161] Reconstitution of a dried blood sample is performed in an extraction buffer that includes a chelating agent, a buffer, and a non-ionic surfactant. In one example, the chelating agent is ethylenediaminetetraacetic acid (EDTA), the buffer is tris(hydroxymethyl) aminomethane (Iris), and surfactant is Triton X-100. The concentrations of EDTA, Tris, and Triton X-100 are selected to both preserve the quality of the genomic DNA and not interfere with downstream library preparation processes, in one example, the extraction buffer is lOmM Tris-HCL pH=8, 10m M EDTA, 0.5% SDS, and 0.25% Triton X-100.

[00162] Extraction of genomic DNA from a reconstituted dried blood spot punch is performed in an extraction buffer at a temperature that maintains the double-stranded nature of the genomic DNA, e.g., below about 90°C or about 60°C. In one example, extraction of genomic DNA from a dried blood spot punch is performed at about 60°C for about 2 hours.

[00163] FIG. 8 illustrates a flow diagram of an example method 800 of extracting genomic DNA from a reconstituted dried blood spot. Method 800 includes, but is not limited to, the following steps:

[00164] In a step 810, dried blood spot (DBS) punches are obtained and placed in individual wells of a microtiter plate. For example, a single 3.2 mm DBS punch for each subject to be tested is obtained and placed in individual wells of a microtiter plate.

[00165] In a step 815, the DBS punches are reconstituted using an extraction buffer. For example, 100 pL of extraction buffer (lOm!V! Tris-HCL pH=8, lOm!V! EDTA, 0.5% SDS, and 0.25% Triton X-100) is added to each well and the plate is incubated for 15 minutes at room temperature. At the end of the incubation period, the supernatant is removed and discarded.

[00166] In a step 820, genomic DNA is extracted in extraction buffer containing proteinase K. For example, 145 pL of extraction buffer (lOmM Tris-HCL pH=^:8, lOmM EDTA, 0.5% SDS, and 0.25%

Triton X-100) and 5 pL of proteinase K are added to each well and the plate is covered and incubated at 60°C with shaking at 1000 rpm for 1 hour ± 15 minutes. At the end of the incubation period, the supernatant from each well is removed to a clean plate (or tubes) and the DBS punch is discarded.

[00167] In a step 825, the extracted genomic DNA is purified for preparation of a sequencing library. For example, the extracted DNA is purified using a magnetic bead-based DNA purification protocol. In one example, 90 pL of Omega Biotech SPRI beads are added to each sample well (or tube) and the reaction is incubated for 2 minutes with mixing. At the end of the incubation period, the beads are washed twice with 80% ethanol and the purified DNA is eluted from the beads using 30 pL of nuclease-free water. [00168] In one aspect, library preparation may be performed using a 96 well microtiter plate format. A 96 well microtiter plate format is widely used and readily available. The 96 well microtiter plate format is also suitable for automation, e.g., for use with liquid-handling robotics.

[00169] FIG. 9 is a table 900 showing the yield metrics for DNA extracted from dried blood spot punches using method 800 of FIG. 8. In this example, statistics for 1,600 1-punch extractions are shown.

[00170] FIG. 10 illustrates a flow diagram of an example method 1000 of preparing a sequencing library from pooled genomic DNA. Method 1000 includes, but is not limited to, the following steps:

[00171] In a step 1010, genomic DNA samples are obtained and pools of individual samples are created. In one example, a two-dimensional pooling strategy of x-number of columns and y-number of rows is used to create z-number of pools. For example, in a 96 well microtiter plate format, 96 different DNA samples extracted in individual wells of a microtiter plate are obtained and a pooling strategy is used to create 20 pools, where 12 "column" or vertical pools each include 8 different samples and 8 "row" or horizontal pools each include 12 different samples. In this pooling strategy, each individual sample is then included in two pools and will ultimately be labeled with 2 different barcodes.

[00172] In a step 1015, the genomic DNA in each pool is fragmented and the fragment ends are repaired. In one example, the genomic DNA is fragmented using an enzymatic fragmentation reaction. The ends of the fragmented DNA are then repaired (e.g., using a blunt ending and dA- tailing reaction) for subsequent ligation to adapter primers.

[00173] In a step 1020, adapter primers are ligated to the fragmented and repaired DNA fragments. In one example, the adapter primers include a hairpin loop that includes a uracil residue to minimize formation of adapter primer dimers. In one example, the adapter primers are NEBNext Adapters obtained from New England BioLabs.

[00174] In a step 1025, the adapter primer ligated DNA is linearized using an enzymatic reaction (e.g., a USER enzyme reaction) to cleave the uracil base in the loop of the adapter primer. The linearized adapter primer ligated DNA is now ready for subsequent hybridization and incorporation of pool-specific "barcode" primers.

[00175] In a step 1030, pool-specific barcode primers are hybridized to the linearized adapter primer ligated DNA. In one example, a pool-specific barcode primer includes a sequence that is complementary to a portion of the adapter primer (for hybridization), a unique barcode sequence that will be used to identify a pool, and sequences for paired end sequencing (e.g., P5 and P7 sequences) on the lllumina MiniSeq SBS platform. In the two-dimensional pooling strategy, each individual subject DNA sample is labeled with two separate pool-level barcodes. A positive "hit" for a variant in an individual subject sample can be identified at the "intersection" of the pools as described with reference to FIG. 4B and FIG. 5. In one example, unique barcode primers with known sequences are obtained from New England BioLabs. In another example, unique barcode primers with known sequences are obtained from Roche Diagnostics. For example, methods of the invention make use of the captured probes target regions (also referred to herein as "markers") identified in Table 2 and Table 3 (below).

[00176] In a step 1035, the adapter ligated and hybridized DNA fragments are PCR amplified to generate pools of uniquely barcoded DNA fragments.

[00177] In a step 1040, the pools of barcoded DNA fragments are combined into a single sample for sequencing.

[00178] In one aspect, a confirmatory test may be used to identify a sample(s) in a positive "hit" pool. In one example, a confirmatory test is a PCR-based test, such as a quantitative PCR (q PCR) test. The qPCR test may, for example, be a multiplexed qPCR test wherein 4 different primer / probe sets with 4 different fluorescent dyes are used in a single reaction to target 4 different variants.

[00179] In one aspect, the confirmatory test (e.g., a multiplexed qPCR test) is performed using extracted genomic DNA obtained from the original reconstituted blood spot extraction protocol.

Variants

[00180] We can precurate known disease specific variants utilizing ClinVar (2018/07/29) as well as manually curated variants from literature review. This will result in an initial version of the Ultra-Low Coverage Sequencing (ULCS) database and will include the variants currently associated with the ULCS panel genes which meet criteria as pathogenic or likely pathogenic variants from multiple submitters in ClinVar as well as manually curated variants. Previously undescribed variants may be categorized as Variants of Unknown Significance (VUS). For example, methods of the invention may be used to screen for any of the disorders or variants identified in Table 1:

Table 1. Screening disorders

Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

1 GAA Pompe Disease (GSD 2) AR 606800 4065 7q25.3

2 IDUA Hurler/Scheie Syndrome (MPS 1) AR 252800 5391 4pl6.3

3 SMN1 Spinal Muscular Atrophy AR 600354 11117 5ql3.2

4 RBI Retinoblastoma, Osteosarcoma AD 180200 9884 13ql4.2

5 ALDOB Hereditary Fructose Intolerance AR 612724 417 9q31.1 Phenotype

Inherit-

Id Gene Disorders MIM HGNC Location ance number

6 ALPL Hypophosphatasia, infantile AR 241500 438 lp36.12

7 ARSA Metachromatic leukodystrophy AR 250100 713 22ql3.33

8 ARSB Maroteaux-Lamy Syndrome (MPS 6) AR 611542 714 5ql4.1

9 ASAH1 Farber lipogranulomatosis and Spinal AR 228000, 735 8p22 muscular atrophy with progressive 159950 myoclonic epilepsy

10 ATP7A Menkes Disease XLR 300011 869 Xq21.1

11 ATP7B Wilson Disease AR 606882 870 13ql4.3

12 C0L3A1 Ehlers-Danlos syndrome, vascular type AD 130050 2201 2q32.2

13 CTNS Cystinosis AR 606272 2518 17pl3.3

14 CYP27A1 Cerebrotendinous xanthomatosis AR 213700 2605 2q35

15 DHCR7 Smith-Lemli-Opitz syndrome AR 270400 2860 llql3.4

16 DMD Duchenne muscular dystrophy XLR 310200 2928 Xp21.2- p21.1

17 GAMT Guanidinoacetate Methyltransferase AR 612736 4136 19ql3.3

18 GJB2 Deafness AR/AD 601544, 4284 13ql2.ll 220290

19 GLA Fabry Disease XLR 300644 4296 Xq22.11

20 GLB1 Morquio B Syndrome (MPS 4B) AR 611458 4298 3p22.3 21 GUSB Sly Disease (MPS 7) AR 611499 4696 7qll.21 22 LIPA Wolman disease AR 278000 6617 10q23.31

23 MAN2B1 Mannosidosis, alpha-, types I and II AR 248500 6826 19pl3.13

24 NAGLU Mucopolysaccharidosis type NIB AR 252920 7632 17q21.2

(Sanfilippo B)

25 NPC1 Niemann-Pick Type C1,D AR 607623 7897 18qll.2

26 OTC Flyperammonemia XLR 311250 8512 Xp21.1

27 SERPINA1 Emphysema/Infantile Cirrhosis AR 107400 8941 14q32.13

28 SGSFI Mucopolysaccharidosis type IIIA AR 252900 10818 17q25.3

(Sanfilippo A)

29 SLC7A7 Lysinuric Protein Intolerance AR 603593 11065 14qll.2

30 SMPD1 Niemann-Pick Disease A, B AR 607608 11120 llpl5.3

31 TPP1 Ceroid lipofuscinosis, neuronal, 2 AR 204500 2073 llpl5.4

32 CYP11B1 Congenital Adrenal Hyperplasia 4 AR 610613 2591 8p21

33 CYP17A1 Congenital Adrenal Hyperplasia 5 AR 202110 2593 10q24.3

34 CYP21A2 Congenital Adrenal Hyperplasia 3 AR 613815 2600 6p21.3

35 DUOX2 Congenital Flypothyroidism AR 606759 13273 15q21.1

(Dysmorphogenesis 6)

36 FISD3B2 Congenital Adrenal Hyperplasia II AR 201810 5218 lpl3.1

37 PAX8 Congenital Flypothyroidism (Thyroid AD 167415 8622 2ql3

Dysgenesis)

38 SLC5A5 Congenital Flypothyroidism AR 601843 11040 19pl3.2-

(Dysmorphogenesis 1) Pl2

39 TG Congenital Flypothyroidism AR 188450 11764 8q24.22

(Dyshormonogenesis 3)

40 TPO Congenital Flypothyroidism AR 606765 12015 2p25.3

(Dyshormonogenesis 2A) Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

41 TSHB Congenital Hypothyroidism (Non- AR 188540 12372 lpl3

Goitrous 4)

42 TSHR Congenital Hypothyroidism (Non- AR 603372 12373 14q31.1

Goitrous)

43 HBB Sickle Cell Disease AR 141900 4827 llpl5.4

44 MTR Methylmalonic/Homocystinuria, cbIG AR 156570 7468 lq43

45 TCN2 Methylmalonic AR 275350 11653 22ql2.2

Aciduria/Homocystinuria Megaloblastic Anemia

46 ASL Argininosuccinic Aciduria AR 207900 746 7qll.21

47 ASS1 Citrullinemia Type 1 AR 215700 758 9q34.13

48 BCKDHA Maple Syrup Urine Disease Type 1A AR 248600 986 19ql3.2

49 BCKDHB Maple Syrup Urine Disease Type IB AR 248600 987 6p22-p21

50 DBT Maple Syrup Urine Disease Type 2 AR 248600 2698 lp31

51 FAH Hepatorenal Tyrosinemia Type 1 AR 276700 3579 15q25.1

52 PAH Phenylketonuria AR 261600 8528 12q23.2

53 ACADM Medium-chain acyl-CoA dehydrogenase AR 201450 89 lp31.1

(MCAD)/Non-Ketotic Hypoglycemia

54 ACADVL Non-Ketotic Hypoglycemia AR 201470 92 17pl3.1

55 HADHA Long-chain 3-hydroxyacyl-CoA AR 143450 4801 2p23 dehydrogenase (LCHAD) deficiency i/Trifunctional protein - _ Subunit

56 HADHB Trifunctional protein - _ Subunit AR 143450 4803 2p23

57 SLC22A5 Primary Systemic Carnitine Deficiency AR 212140 10969 5q31.1

58 MMADHC Methylmalonic/Homocystinuria, cbID AR 277410 25221 2q23.2

59 ACAT1 a-Methylacetoacetic Aciduria 1 ( b - AR 203750 93 llq22.3

Ketothiolase Deficiency)

60 GCDH Glutaric Aciduria Type 1 AR 231670 4189 19pl3.2 61 HMGCL 3-Hydroxy-3-Methylglutaric Aciduria AR 246450 5005 lp36.1-

Ketoacidosis p35

62 IVD Isovaleric Acidemia AR 243500 6186 15ql4- ql5

63 MCCC1 3-Methylcrotonylglycinuria 1 AR 210200 6936 3q25-q27

64 MCCC2 3-Methylcrotonylglycinuria 2 AR 609014 6937 5ql2-ql3

65 MCEE Methylmalonic Aciduria, Mut Type AR 608419 16732 2pl3.3

66 MMAA Methylmalonic Aciduria, cblA Type AR 251100 18871 4q31.21

67 MMAB Methylmalonic Aciduria, cbIB Type AR 251110 19331 12q24

68 MMUT Methylmalonic Aciduria, Mut Type AR 251000 7526 6p21.2- p21.1

69 PCCA Propionic Aciduria Type 1 AR 282000 8653 13q32.3

70 PCCB Propionic Aciduria Type 2 AR 532000 8654 3q22.3

71 ABCD1 Adrenoleukodystrophy XLR 300371 61 Xq28

72 BTD BTD-Late Onset Multiple Carboxylase AR 253260 1122 3p25

73 CFTR Cystic Fibrosis AR 602421 1884 7q31.2

74 GALT Galactosemia 1 AR 606999 4135 9pl3.3

75 IL2RG Severe Combined Immunodeficiency XLR 308380 6010 Xql3.1 Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

76 JAK3 Severe Combined Immunodeficiency AR 600173 6193 19pl3.ll

77 ZAP70 Severe Combined Immunodeficiency AR 176947 12858 2qll.2

78 ADA Severe Combined Immunodeficiency AR 102700 186 20ql2- ql3.1

79 CBS Homocystinuria, B6-responsive and AR 236200 1550 21q22.3 nonresponsive types

80 CD3D Severe Combined Immunodeficiency AR 615617 1673 llq23.3

81 CD3E Severe Combined Immunodeficiency AR 615615 1674 llq23.3

82 CPS1 Hyperammonemia AR 237300 2323 2q34

83 DCLRE1C Severe Combined Immunodeficiency AR 605988 17642 10pl3

84 DLD Maple Syrup Urine Disease Type 3 AR 246900 2898 7q31-q32

85 DNAJC12 Hyperphenylalaninemia (non-BH4- AR 617384 28908 10q21.3 deficient)

86 DUOXA2 Congenital Hypothyroidism AR 612772 32698 15q21.1

(Dysmorphogenesis 5)

87 FOXE1 Congenital Hypothyroidism (Bamforth- AR 602617 3806 9q22.33

Lazarus Syndrome)

88 G6PD Hemolytic Anemia XLR 305900 4057 Xq28

89 GALC Krabbe Disease AR 606890 4115 14q31

90 GALNS Morquio A Syndrome (MPS 4A) AR 612222 4122 16q24.3

91 GATM CCDS AR 612718 4175 15q21.1

92 GBA Gaucher Disease AR 606463 4177 lq22

93 GCH1 Hyperphenylalaninemia (Biopterin AD 233910 4193 14q22.2

Cofactor Defect B)

94 GSS 5-Oxoprolinuria (Pyroglutamic Aciduria) AR 266130 4624 20qll.21

95 HLCS Multiple CoA Carboxylase Deficiency AR 253270 4976 21q22.2

(Biotin Responsive)

96 IDS Hunter Syndrome (MPS 2) XLR 300823 5389 Xq28

97 IL2RA Severe Combined Immunodeficiency AR 606367 6008 10pl5.1

98 IL7R Severe Combined Immunodeficiency AR 608971 6024 5pl3.2

99 IYD Congenital Hypothyroidism AR 612025 21071 6q25.1

100 LCK Severe Combined Immunodeficiency AR 615758 6524 lp35.1

101 LHX3 Congenital Hypothyroidism (Combined AR 600577 6595 9q34.3

Pituitary Deficiency 3)

102 LIG4 Severe Combined Immunodeficiency AR 601837 6601 13q33.3

103 MTHFR Homocystinuria due to MTHFR AR 236250 7436 lp36.22 deficiency

104 MTRR Homocystinuria-megaloblastic anemia, AR 236270 7473 5pl5.31 cbl E type

105 NHEJ1 Severe Combined Immunodeficiency AR 311290 25737 2q35

106 NKX2-1 Congenital Hypothyroidism (Goitrous) AD 600635 11825 14ql3.3

107 NKX2-5 Congenital Hypothyroidism (Non- AD 600584 2488 5q35.1

Goitrous 5)

108 NR0B1 Congenital Adrenal Hyperplasia XLR 300473 7960 Xp21.2

109 PCBD1 Hyperphenylalaninemia (Biopterin AR 264070 8646 10q22

Cofactor Defect D) Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

110 PNP Severe Combined Immunodeficiency AR 613179 7892 14qll.2

111 POU1F1 Congenital Flypothyroidism (Combined AR 173110 9210 3pll.2 Pituitary Flormone Deficiency)

112 PPM1K Maple Syrup Urine Disease (Mild) 615135 25415 4q22.1

113 PROP1 Congenital Flypothyroidism (Combined AR 601538 9455 5qll.2 Pituitary Flormone Deficiency)

114 PTPRC Severe Combined Immunodeficiency AR 608971 9666 lq31.3

115 PTS Flyperphenylalaninemia (Biopterin AR 261640 9689 llq23.1 Cofactor Defect A)

116 QDPR Flyperphenylalaninemia (Biopterin AR 261630 9752 4pl5.32 Cofactor Defect C)

117 RAG1 Severe Combined Immunodeficiency AR 179615 9831 llpl2 (Omenn Syndrome)

118 RAG 2 Severe Combined Immunodeficiency AR 179616 9832 llpl2 (Omenn Syndrome)

119 SLC25A15 HHH Syndrome AR 238970 10985 13ql4.ll

120 SPR Flyperphenylalaninemia(Biopterin AR 612716 11257 2pl4-pl2 Cofactor Defect)

121 STAR Congenital Lipoid Adrenal Hyperplasia I AR 600617 11359 8pll.23

122 SUCLA2 Mitochondrial DNA Depletion Syn 5 AR 603921 11448 13ql4.2 (Methylmalonic Aciduria)

123 SUCLG1 Mitochondrial DNA Depletion Syn 9 AR 611224 11449 2pll.2 (Methylmalonic Aciduria)

124 THRA Congenital Flypothyroidism (Non- AD 190120 11796 17q21.1 Goitrous 6)

125 TRHR Congenital Flypothyroidism AR 188545 12299 8q23.1

126 AHCY Flypermethioninemia AR 613752 343 20qll.22

127 ARG1 Argininemia AR 207800 663 6q23.2

128 GNMT Flypermethioninemia AR 606664 4415 6p21.1

129 HPD Flereditary Tyrosinemia Type 3 AR 276710 5147 12q24.31

(Flawkinsinuria)

130 MAT1A Methionine adenosyltransferase AR/AD 250850 6903 10q22 deficiency

131 SLC25A13 Citrullinemia Type 2 AR 605814 10983 7q21.3

132 TAT Oculocutaneous Tyrosinemia ( AR 276600 11573 16q22.2 TyrosinemiaType 2)

133 CPT1A Non-Ketotic Flypoglycemia AR 255120 2328 llql3.2

134 CPT2 Non-Ketotic Flypoglycemia; Myopathy AR 255110 2330 lp32.3

135 DECR1 Non-Ketotic Flypoglycemia AR 222745 2753 8q21.3

136 ETFA Glutaric Acidemia Type 2A AR 608053 3481 15q24.2

137 ETFB Glutaric Acidemia Type 2B AR 130410 3482 19ql3.33

138 ETFDH Glutaric Acidemia Type 2C AR 231675 3483 4q32.1

139 HADH Flyperinsulinemic Flypoglycemia 4 AR 601609 4799 4q22-q26

140 SLC25A20 Carnitine Translocase Deficiency AR 613698 1421 3p21.31

141 ABCD4 Methylmalonic/Flomocystinuria CblJ AR 603214 68 14q24.3

142 ACAD8 Isobutyryl-CoA Dehydrogenase AR 604773 87 llq25 Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

143 ACADSB 2-Methylbutyryl Glycinuria AR 600301 91 10q26.13

144 AUH 3-Methylglutaconic Aciduria Type 1 AR 250950 890 9p22

145 HCFC1 Methylmalonic/Homocystinuria XLR 309541 28908 10q21.3

146 HSD17B10 17- ?-OH-Steroid Dehydrogenase Def XLD 300256 4800 Xpll.22

147 LMBRD1 Methylmalonic/Homocystinuria, cblF AR 277380 23038 6ql3

148 MMACHC Methylmalonic/Homocystinuria, cbIC AR 277400 24525 lp34.1

149 GALE Galactosemia AR 230350 4116 Ip36-p35

150 GALK1 Galactosemia AR 604313 4118 17q24

151 ACADS Non-Ketotic Hypoglycemia AR 201470 90 12q24.31

152 ADK Hypermethioninemia (Adenosine AR 102750 257 10q22.2

Kinase)

153 DNAJC19 3-Methylglutaconic Aciduria AR 608977 30528 3q26.33

154 MLYCD Malonic Aciduria AR 248360 7150 16q23.3- q24

155 NADK2 2,4 Dienoyl-CoA reductase deficiency AR 616034 26404 5pl3.2

156 OP A3 3-Methylglutaconic Aciduria Type 3 AR/AD 258501 8142 19ql3.32

(Costeff Optic Atrophy Syndrome)

157 SERAC1 3-Methylglutaconic Aciduria AR 614739 21061 6q25.3

158 TAZ 3-Methylglutaconic Aciduria Type 2 XLR 302060 11577 Xq28

(Barth Syndrome)

159 ALDH4A1 Hyperprolinemia Type 2 AR 606811 406 lp36.13

160 AMT Nonketotic Hyperglycinemia 2 AR 238310 473 3p21.2

161 ETHE1 Ethylmalonic Encephalopathy 1 AR 608451 23287 19ql3.31

162 GLDC Nonketotic Hyperglycinemia 1 AR 238300 4313 9p24.1

163 NPC2 Niemann-Pick Type C2 AR 601015 14537 14q24.3

164 OAT Hyperornithinemia (Gyrate Atrophy of AR 258870 8091 10q26.13 Choroid & Retina)

165 OPLAH 5-Oxoprolinuria (Pyroglutamic Aciduria) AR/AD 260005 8149 8q24

166 PRODH Hyperprolinemia Type 1 (Proline AR 606810 9453 22qll.21

Oxidase 1)

167 SLC6A8 Creatine Deficiency Syndrome XLR 300036 11055 Xq28

168 ALK Neuroblastoma 613014 427 2p23.2- p23.1

169 APC familial adenomatous polyposis-1 AD 175100 583 5q22.2 (FAP1), adenomatous polyposis coli

170 DICERl PPB Familial Tumor and Dysplasia AD 606241 17098 14q32.13 Syndrome, DICERl Syndrome

171 PHOX2B Neuroblastoma, Central AD 209880 9143 4pl3 hypoventilation syndrome

172 PTCH1 Nevoid Basal Cell Carcinoma Syndrome AD 109400 9585 9q22.32 (Gorlin Syndrome)

173 RET Hirschsprung Disease I AD 142623 9967 10qll.21

174 SMARCB1 Rhabdoid Tumor Predisposition AD 609322 11103 22qll.23 Syndrome

175 SUFU Medulloblastoma AR/AD 155255 16466 10q24.32

176 TP53 Li-Fraumeni Syndrome AD 151623 11998 17pl3.1 Phenotype

Inherit¬

Id Gene Disorders MIM HGNC Location ance number

177 WT1 Wilms tumor, type 1 and other WT1- AD 194070 12796 llpl3 related disorders

178 ACADL Nonketotic Hypoglycemia AR 609576 88 2q34

179 FTCD Glutamate formiminotransferase AR 229100 3974 21q22.3 deficiency

180 HSD17B4 17- ?-OH-Steroid Dehydrogenase 4 AR 601860 5213 5q23.1

181 MVK Mevalonic Aciduria AR 251170 7530 12q24

182 NAGS Hyperammonemia AR 608300 17996 17q21.31

183 SUGCT Glutaric Aciduria Type 3 AR 231690 16001 7pl4.1

Examples

[00181] DNA extraction and library preparation may be performed from a single 3.2 mm punch based on our previously reported methods (Genetics in Medicine 2015). DNA may be subjected to pooled library preparation for sequencing on the lllumina MiniSeq using genomic enrichment methods modified from the previously described protocol (Genetics in Medicine 2015). Briefly, isolated DNA may be fragmented enzymatically, with DNA ends adaptor-ligated for barcoding and incubated with oligonucleotide probes for hybrid capture followed by next generation sequencing (NGS). For library production, we can utilize the New England BioLabs Inc. NEBNext^® Ultra™ II FS DNA Module (E7810) for DNA fragmentation during a thermal cycler incubation. Library construction may be considered acceptable when >90% pooled samples yield the following criteria: pre-capture library yield >500 ng; pre-Capture library peak size between 215-350 bp; adapter contamination <10% and overamplification products <10%. For optimized sequencing performance, hybridization capture of pooled libraries may be considered acceptable if: post capture single pool yield is >100 ng; post-capture library peak size is between 280-360 bp, amplification primer contamination <10%, overamplification <10%, and <lper-library demultiplex representative of 2-8% of sequencing reads.

[00182] Sequencing can take place as a paired end (2x75bp) run on the lllumina MiniSeq Sequencing by Synthesis (SBS) platform using genomic enrichment methods modified from the previously described protocol(Genetics in Medicine 2015). After sequencing reads are generated, the first step in the analysis may be the "demultiplexing" (assignment of reads to individual pooled samples based on the dual indexes) and generation of FASTQ data files. Picard and other tools may be used to assess sequencing QC and enrichment specific statistics. Next, variant calling may be performed from the FASTQ files using a string search algorithm using a custom data analysis pipeline from a list of pre-defined SNP/lndel variants. Next, we can also call out CNV based on read-depth coverage for a pre-defined list of CNV variants. All QC metrics from the bioinformatic analysis for a clinical sample from custom tools may be aggregated into a single report. Finally, .vcf (variant call files) are generated for each sample and uploaded to a custom portal for reporting.

[00183] To demonstrate proof of principle of using a two-dimensional pooling strategy for variant screening, computer modeling was used to simulate different aspects of the process.

[00184] To simulate variant "hit rate", a comparison of empirical and theoretical estimates were calculated for Pompe's disease. For the "hit" rate comparison, we calculated variant frequency for Pompe's disease which has a known incidence of 1:40,000 individuals. We calculated an empirical estimate using a dataset from an Oregon study, where we know to expect 181 Pompe "hits" out of 40,000 individuals screened, i.e., a "hit rate" of about 0.45% or 1 out of 221. Second tier screens of 599 GAA variants (i.e., GAA gene variants for acid alpha-glucosidase) were subdivided as: 1 positive, 180 indeterminate, 235 pseudodeficiency (negative), and 183 no variants detected (negative). The number of positive Pompe hits is 1 positive plus 180 indeterminate or 181.

[00185] Using the gnomAD and ClinVar datasets, we estimated a theoretical hit rate for Pompe disease based on 217 ClinVar pathogenic (P)/ likely pathogenic (LP) variants modeled using the gnomAD dataset to be about 0.6% or 1 out of 159, which is consistent with the empirical dataset (exceptions are variants of unknown significance (VUS)).

[00186] If variant screening is performed using a two-dimensional pooling strategy, we estimate that about 80% of the second-tier screening samples (i.e., the 599 GAA variants) will be captured and 100% may be captured if variants of unknown significance (VUS) are included in the screening panel (ITT panel).

[00187] For genetic screening, we can think of affected or incidence of 1: 48,800 to 1:25,281 and then determine the carrier frequency.

[00188] FIG. 11 is a diagram 1100 of the empirical and theoretical estimates for carrier frequency.

[00189] Modeling was also performed to simulate screening for a low frequency variant and a high frequency variant using a two-dimensional pooling strategy and define a point (cutoff) at which a variant hit will be resolvable (i.e., at the sample level) or unresolvable (i.e., at the pool level that will require second level testing).

[00190] FIG. 12 is a plot 1200 showing hits identified in a two-dimensional pooling strategy for a low frequency variant (VI) and a high frequency variant (V2). In this example, 100 samples were divided into 2 sets of pools, i.e., 10 pools in the horizontal dimension (pools 1 through 10) and 10 pools in the vertical dimension (pools 11 through 20), wherein each pool included 10 samples. Positive pools identified were pools 1, 5, 7, 12, 14, and 17, shaded light gray. The low frequency variant VI was found in one sample identified by the intersection of the positive pools 7 and 14 (i.e., a correct hit), shaded dark gray. The high frequency variant V2 was identified by the intersection of positive pools 1 and 12; positive pools 1 and 17; positive pools 5 and 12; and positive pools 5 and 17. These pools are shaded medium gray. The simulation shows that DNA pools with a low frequency variant are resolvable to the sample level, while DNA pools with a high frequency variant have overlapping, unresolvable result (i.e., incorrect result) that will require a second level of testing to resolve.

[00191] In one embodiment, second level testing using targeted next-generation sequencing or PCR may be used to resolve which sample has the positive result. In one aspect, a unique barcode is added to every ambiguous sample and then each ambiguous sample is tested individually. In another aspect, when there is a positive result on a row of the matrix but no corresponding positive result on the vertical column of the matrix, the samples from the row of the matrix are separated into a smaller pool where individual barcodes are coupled to each sample in this new set and the resulting new data set is sequenced in a one-dimensional manner. In this aspect, the entire row of the matrix of samples containing the ambiguous samples are tested individually. In another aspect, when there is a positive result on a column of the matrix but no corresponding positive result on the horizontal row of the matrix, the samples from the column of the matrix are separated into a smaller pool where individual barcodes are coupled to each sample in this new set and the resulting new data set is sequenced in a one-dimensional manner. In this aspect, the entire column of the matrix of samples containing the ambiguous samples are tested individually.

[00192] Rare variants are unlikely to create errors using a two-dimensional screening strategy compared to high frequency variants. To further demonstrate the effect of variant frequency on the number of incorrect hits in a two-dimensional screening strategy, a simulated screening of 100,000 newborns for Pompe variants was performed.

[00193] FIG. 13 is a Table 1300 showing the computer simulated two-dimensional screening of 100,000 newborns based on GAA variant frequency. In this example, two different target panels based on variant frequency were used: a rare GAA pathogenic (P)/ likely pathogenic (LP) variant panel and a common GAA pseudodeficiency variant plus pathogenic (P)/ likely pathogenic (LP) variant panel. The GAA P/LP variants in the rare variant panel have an aggregate frequency of about 0.5%, which is about 1 in 200 individuals. The aggregate frequency for the common variant panel (i.e., GAA pseudodeficiency + P/LP) is about 8.5%, which is about 1 in 12 individuals. The higher aggregate frequency for the common variant panel is due to the inclusion of pseudodeficiency variants that are relatively common in the population. The simulated screening of the 100,000 samples for the two different target panels was done as 1,000 runs at 100 samples/run. The data show that when screening 100,000 newborns for the rare variant panel (i.e., GAA P/LP), the number of "incorrect" hit samples (or "unresolvable" samples) is about 97 (about 0.1% or less unresolvable). For the common variant panel (i.e., GAA pseudodeficiency + P/LP), the number of incorrect hit samples (or unresolvable samples) is relatively high at about 19447 (about 20% unresolvable).

[00194] FIG. 14A is a plot 1400 of the frequency of incorrect hits per run for the rare variant panel of FIG. 13. Plot 1400 shows the number of times incorrect "hit" or errors occurred in the run. The data show that for the rare variant panel the majority of the runs have zero errors and can be resolved.

[00195] FIG. 14B is a plot 1410 of the common variant panel of FIG. 13. Plot 1410 shows the number of times incorrect "hits" or errors occurred in the run.

[00196] FIG. 15 illustrates a schematic diagram 1500 of the expected and experimentally observed hits for a high frequency variant GAA c.-32-13T>G in a two-dimensional screening. In the 10 X 10 two-dimensional screening, "0" represents a negative (Neg) and "1" represents a positive for the high frequency variant. For the high frequency variant (GAA c.-32-13T>G), we expect to see three hits as shown in the "Expected" panel on the top. Experimentally with a two-dimensional pooling strategy, 6 incorrect hits (unresolvable cases) for the high frequency variant (GAA c.-32-13T>G) are observed.

[00197] To demonstrate the feasibility of detecting a positive at the pool level using a one dimensional pooling strategy, 2 sets of n=10 and n=20 pooled samples were used. A first set of pooled samples included 1 sample that was positive for a P/LP variant (i.e., GAA c. 1979G>A) for Pompe disease and the rest of the samples were negative; and a second set of pooled samples included 1 sample that was positive for 2 pseudodeficiency variants (i.e., GAA c. 1726G>A and GAA c. 2065G>A) for Pompe disease and the rest of the samples were negative.

[00198] FIG. 16A and FIG. 16B illustrate schematic diagrams 1600 and 1610 of a one-dimensional screening strategy for detecting known GAA variants in 2 sets of pooled samples. In this example, each set of pooled samples includes a pool of n=10 samples and a pool of n=20 samples. Each sample pool in the set is represented in duplicate (i.e., n=2, "Rep 1" and "Rep 2").

[00199] With reference to FIG. 16A, a first set of pooled samples includes a single sample with a Pompe P/LP variant GAA c. 1979G>A.

[00200] With reference to FIG. 16B, a second set of pooled samples includes a single sample with 2 pseudodeficiency variants GAA c. 1726G>A and GAA c. 2065G>A. [00201] FIG. 17A is a plot 1700 of expected variant coverage in the first set of n = 10 unpooled samples of FIG. 16A versus observed variant coverage in the pooled samples. The arrow indicates the known P/LP variant GAA c. 1979G>A.

[00202] FIG. 17B is a plot 1710 of expected variant coverage in the first set of n = 20 unpooled samples of FIG 16A versus observed variant coverage in the pooled samples. The arrow indicates the known P/LP variant GAA c. 1979G>A.

[00203] With reference to FIG. 17A and FIG. 17B, the data show that the Pompe disease causing P/LP variant GAA c. 1979G>A was successfully detected using a string search algorithm for SNV in both n = 10 and n = 20 pooled DNA samples. As the DNA pool size increased from n = 10 to n = 20, an increase in "variant dropout" (i.e., the chances of a variant being covered decreases as the amount of genomic DNA increases) was observed (indicated by the boxed region on each plot). The data also shows that we were able to detect insertions and deletions (INDELs).

[00204] FIG. 18A is a plot 1800 of expected variant coverage in the second set of n = 10 unpooled samples of FIG. 16B versus observed variant coverage in the pooled samples. The arrows indicate the known pseudodeficiency variants GAA c. 1726G>A and GAA c. 2065G>A.

[00205] FIG. 18B is a plot 1800 of expected variant coverage in the second set of n = 20 unpooled samples of FIG. 16B versus observed variant coverage in the pooled samples. The arrows indicate the known pseudodeficiency variants GAA c. 1726G>A and GAA c. 2065G>A.

[00206] With reference to FIG. 18A and FIG. 18B, the data show that the pseudodeficiency variants GAA c. 1726G>A and GAA c. 2065G>A were successfully detected using a string search algorithm for SNV in both n = 10 and n = 20 pooled DNA samples. As the DNA pool size increased from n = 10 to n = 20, an increase in variant dropout was again observed. Insertions and deletions (INDELs) were also detected.

[00207] FIG. 19A is a plot 1900 of a comparison of variant coverage within the n = 10 DNA pooled experimental replicates of FIG. 16A. In the n = 10 DNA pooled experimental replicates there were 9 dropout variants. The data show that some of the variants were covered in pool n = 10 replicate 1 (Pool 10 Repl) and some of the variants were covered in pool n = 10 replicate 2 (Pool 10 Rep2).

[00208] FIG. 19B is a plot 1910 of a comparison of variant coverage within the n = 20 DNA pooled experimental replicates of FIG. 16A. The data show that in the n = 20 DNA pooled experimental replicates there were more variant dropouts compared to the n = 10 DNA pooled replicates of FIG. 18A. Some of the variants were covered in pool n = 20 replicate 1 (Pool 20 Repl) and some of the variants were covered in pool n = 20 replicate 2 (Pool 20 Rep2). The variant dropout effect in a DNA pool of 20 samples may, for example, be improved by reducing the sequencing target panel size and/or adjusting experimental conditions (e.g., reagent concentrations).

[00209] FIG. 19C is a plot 1920 of a comparison of variant dropouts between the n = 10 and n = 20 DNA pooled experimental replicates of FIG. 16A.

Table 2. Target regions for the PD4111 Gene Newborn Screening Panel

Chromosome Beginning End Condition chrl 11850728 11850968 MTHFR chrl 11851218 11851378 MTHFR chrl 11852281 11852481 MTHFR chrl 11853906 11854186 MTHFR chrl 11854400 11854640 MTHFR chrl 11854734 11854974 MTHFR chrl 11855115 11855423 MTHFR chrl 11856219 11856499 MTHFR chrl 11860208 11860408 MTHFR chrl 11861221 11861461 MTHFR chrl 11862936 11863216 MTHFR chrl 21880535 21880695 ALPL chrl 21887086 21887286 ALPL chrl 21887548 21887748 ALPL chrl 21889547 21889787 ALPL chrl 21890488 21890768 ALPL chrl 21894549 21894789 ALPL chrl 21896761 21896921 ALPL chrl 21900107 21900347 ALPL chrl 21902178 21902458 ALPL chrl 21902976 21903176 ALPL chrl 21903871 21904151 ALPL chrl 24122608 24122808 GALE chrl 24122977 24123350 GALE chrl 24123518 24123718 GALE chrl 24124157 24124357 GALE chrl 24124570 24124770 GALE chrl 24125083 24125243 GALE chrl 24125298 24125498 GALE chrl 24128915 24129075 HMGCL chrl 24130853 24131013 HMGCL chrl 24134626 24134866 HMGCL chrl 24137174 24137374 HMGCL chrl 24140624 24140824 HMGCL chrl 24143127 24143327 HMGCL chrl 24143903 24144103 HMGCL chrl 24146918 24147118 HMGCL chrl 24151794 24151954 HMGCL chrl 32745229 32745785 LCK chrl 76190386 76190586 ACADM chrl 76194058 76194218 ACADM chrl 76198278 76198478 ACADM chrl 76198493 76198653 ACADM chrl 76199163 76199363 ACADM chrl 76200440 76200600 ACADM chrl 76205610 76205850 ACADM chrl 76211407 76211647 ACADM Chromosome Beginning End Condition chrl 76215054 76215294 ACADM chrl 76216079 76216279 ACADM chrl 76226789 76227069 ACADM chrl 76228327 76228487 ACADM chrl 100661726 100662006 DBT chrl 100671677 100671877 DBT chrl 100671991 100672231 DBT chrl 100676149 100676349 DBT chrl 100680317 100680597 DBT chrl 100681535 100681775 DBT chrl 100684081 100684353 DBT chrl 100696236 100696516 DBT chrl 100700944 100701144 DBT chrl 100706304 100706464 DBT chrl 100715225 100715425 DBT chrl 115576034 115576194 TSHB chrl 115576536 115576904 TSHB chrl 119957974 119958174 HSD3B2 chrl 119962090 119962250 HSD3B2 chrl 119964490 119965010 HSD3B2 chrl 119965085 119965285 HSD3B2 chrl 155204784 155205104 GBA chrl 155205413 155205693 GBA chrl 155205998 155206278 GBA chrl 155207139 155207379 GBA chrl 155207873 155208153 GBA chrl 155208256 155208496 GBA chrl 155209389 155209549 GBA chrl 155209674 155209874 GBA chrl 155210362 155210522 GBA chrl 198671560 198671760 PTPRC chrl 198685870 198686070 PTPRC chrl 198687296 198687496 PTPRC chrl 198704232 198704432 PTPRC chrl 236958905 236959105 MTR chrl 236969343 236969543 MTR chrl 236971949 236972149 MTR chrl 236987420 236987620 MTR chrl 236988600 236988800 MTR chrl 236998786 236998986 MTR chrl 237013589 237013789 MTR chrl 237015778 237015978 MTR chrl 237023077 237023277 MTR chrl 237024403 237024603 MTR chrl 237026653 237026853 MTR chrl 237038014 237038214 MTR chrl 237044013 237044213 MTR chrl 237052500 237052700 MTR chrl 237057731 237057931 MTR chrl 237058651 237058811 MTR chrl 237060214 237060414 MTR chr2 1439947 1440147 TPO chr2 1459889 1460049 TPO chr2 1481122 1481322 TPO chr2 1488277 1488656 TPO chr2 1491513 1491863 TPO Chromosome Beginning End Condition chr2 1497509 1497854 TPO chr2 1499731 1499931 TPO chr2 1500361 1500521 TPO chr2 1507686 1507886 TPO chr2 26414083 26414243 HADHA chr2 26414291 26414531 HADHA chr2 26415121 26415321 HADHA chr2 26416432 26416672 HADHA chr2 26417384 26417544 HADHA chr2 26417904 26418104 HADHA chr2 26420506 26420666 HADHA chr2 26423966 26424282 HADHA chr2 26426909 26427109 HADHA chr2 26432596 26432796 HADHA chr2 26435392 26435552 HADHA chr2 26437315 26437475 HADHA chr2 26437932 26438132 HADHA chr2 26453011 26453211 HADHA chr2 26454974 26455174 HADHA chr2 26457034 26457274 HADHA chr2 26459678 26459878 HADHA chr2 26461745 26461905 HADHA chr2 26461907 26462107 HADHA chr2 26467353 26467513 HADHA chr2 26477010 26477210 HADHB chr2 26486234 26486434 HADHB chr2 26492763 26492923 HADHB chr2 26496417 26496617 HADHB chr2 26499843 26500043 HADHB chr2 26501501 26501701 HADHB chr2 26501961 26502365 HADHB chr2 26502842 26503042 HADHB chr2 26505816 26506016 HADHB chr2 26507694 26507894 HADHB chr2 26508267 26508467 HADHB chr2 26512684 26512884 HADHB chr2 71351475 71351675 MCEE chr2 98340402 98340840 ZAP70 chr2 98349256 98349456 ZAP70 chr2 98349659 98350142 ZAP70 chr2 98351072 98351272 ZAP70 chr2 98351794 98351994 ZAP70 chr2 98353940 98354140 ZAP70 chr2 98354189 98354349 ZAP70 chr2 98354398 98354598 ZAP70 chr2 98355749 98355949 ZAP70 chr2 113984684 113984884 PAX8 chr2 113992958 113993158 PAX8 chr2 113999147 113999347 PAX8 chr2 114001971 114002171 PAX8 chr2 114004284 114004484 PAX8 chr2 150426518 150426718 MMADHC chr2 150427495 150427695 MMADHC chr2 150432184 150432384 MMADHC chr2 150432904 150433064 MMADHC chr2 150435935 150436215 MMADHC Chromosome Beginning End Condition chr2 150438593 150438833 MMADHC chr2 189839145 189839345 COL3A1 chr2 189849438 189849789 COL3A1 chr2 189850347 189850547 COL3A1 chr2 189851763 189851923 COL3A1 chr2 189852761 189852921 COL3A1 chr2 189853266 189853426 COL3A1 chr2 189854070 189854230 COL3A1 chr2 189854764 189854924 COL3A1 chr2 189854981 189855141 COL3A1 chr2 189855678 189855838 COL3A1 chr2 189856154 189856505 COL3A1 chr2 189856854 189857014 COL3A1 chr2 189857546 189857706 COL3A1 chr2 189858039 189858239 COL3A1 chr2 189858716 189858876 COL3A1 chr2 189858906 189859106 COL3A1 chr2 189859215 189859375 COL3A1 chr2 189859412 189859572 COL3A1 chr2 189859713 189859873 COL3A1 chr2 189860384 189860544 COL3A1 chr2 189860798 189860958 COL3A1 chr2 189861081 189861281 COL3A1 chr2 189861833 189861993 COL3A1 chr2 189862011 189862171 COL3A1 chr2 189862373 189862533 COL3A1 chr2 189862942 189863102 COL3A1 chr2 189863339 189863499 COL3A1 chr2 189863997 189864317 COL3A1 chr2 189864516 189864676 COL3A1 chr2 189866096 189866376 COL3A1 chr2 189866970 189867130 COL3A1 chr2 189867626 189867826 COL3A1 chr2 189868086 189868246 COL3A1 chr2 189868409 189868569 COL3A1 chr2 189868677 189868917 COL3A1 chr2 189868937 189869137 COL3A1 chr2 189870033 189870233 COL3A1 chr2 189870932 189871172 COL3A1 chr2 189871579 189871739 COL3A1 chr2 189872177 189872377 COL3A1 chr2 189872595 189872875 COL3A1 chr2 189873598 189873758 COL3A1 chr2 189873848 189874048 COL3A1 chr2 189874828 189875149 COL3A1 chr2 189875354 189875634 COL3A1 chr2 189876312 189876552 COL3A1 chr2 211342389 211342589 CPS1 chr2 211421371 211421684 CPS1 chr2 211437979 211438179 CPS1 chr2 211441025 211441265 CPS1 chr2 211444392 211444552 CPS1 chr2 211447291 211447491 CPS1 chr2 211452772 211452972 CPS1 chr2 211454774 211455014 CPS1 chr2 211456552 211456752 CPS1 Chromosome Beginning End Condition chr2 211457513 211457713 CPS1 chr2 211459195 211459395 CPS1 chr2 211460190 211460350 CPS1 chr2 211464048 211464364 CPS1 chr2 211465248 211465448 CPS1 chr2 211466914 211467114 CPS1 chr2 211469828 211469988 CPS1 chr2 211471412 211471692 CPS1 chr2 211473001 211473338 CPS1 chr2 211476799 211477039 CPS1 chr2 211481074 211481274 CPS1 chr2 211502429 211502589 CPS1 chr2 211503834 211503994 CPS1 chr2 211504671 211504871 CPS1 chr2 211507130 211507410 CPS1 chr2 211512570 211512770 CPS1 chr2 211513132 211513332 CPS1 chr2 211515046 211515246 CPS1 chr2 211518711 211518871 CPS1 chr2 211521195 211521355 CPS1 chr2 211523263 211523463 CPS1 chr2 211525127 211525461 CPS1 chr2 211527811 211528011 CPS1 chr2 211532882 211533082 CPS1 chr2 211539536 211539736 CPS1 chr2 211540466 211540666 CPS1 chr2 211541714 211541914 CPS1 chr2 219646861 219647198 CYP27A1 chr2 219674255 219674535 CYP27A1 chr2 219676920 219677200 CYP27A1 chr2 219677234 219677514 CYP27A1 chr2 219677592 219677872 CYP27A1 chr2 219678688 219679006 CYP27A1 chr2 219679091 219679451 CYP27A1 chr2 219679590 219679790 CYP27A1 chr2 219942822 219942982 NHEJ1 chr2 220011360 220011520 NHEJ1 chr2 220022147 220022307 NHEJ1 chr2 220022811 220023173 NHEJ1 chr3 15643301 15643461 HACL1 chr3 15676970 15677210 BTD chr3 15683369 15683609 BTD chr3 15685807 15687007 BTD chr3 87309089 87309289 P0U1F1 chr3 87310373 87310533 P0U1F1 chr3 87311200 87311400 P0U1F1 chr3 87313359 87313559 P0U1F1 chr3 87313565 87313765 P0U1F1 chr3 87322532 87322732 P0U1F1 chr3 87325442 87325642 P0U1F1 chr3 135969125 135969457 PCCB chr3 135974602 135974842 PCCB chr3 135975358 135975518 PCCB chr3 135979276 135979436 PCCB chr3 135980771 135980931 PCCB chr3 136002634 136002834 PCCB Chromosome Beginning End Condition chr3 136003097 136003297 PCCB chr3 136012567 136012767 PCCB chr3 136016734 136016974 PCCB chr3 136019812 136020012 PCCB chr3 136035691 136036008 PCCB chr3 136045580 136045780 PCCB chr3 136045952 136046152 PCCB chr3 136046425 136046625 PCCB chr3 136047586 136047746 PCCB chr3 136048702 136048902 PCCB chr3 182733223 182733383 MCCC1 chr3 182737840 182738040 MCCC1 chr3 182740193 182740393 MCCC1 chr3 182743495 182743695 MCCC1 chr3 182751724 182751924 MCCC1 chr3 182754962 182755320 MCCC1 chr3 182756776 182756976 MCCC1 chr3 182759314 182759554 MCCC1 chr3 182763160 182763360 MCCC1 chr3 182769857 182770057 MCCC1 chr3 182775046 182775206 MCCC1 chr3 182788803 182789083 MCCC1 chr3 182804394 182804594 MCCC1 chr3 182810220 182810380 MCCC1 chr3 182812294 182812494 MCCC1 chr3 182817084 182817284 MCCC1 chr4 980784 981131 IDUA chr4 981516 981796 IDUA chr4 983554 983754 IDUA chr4 984838 985038 IDUA chr4 994343 994543 IDUA chr4 994623 994823 IDUA chr4 995204 995404 IDUA chr4 995442 995682 IDUA chr4 995720 996000 IDUA chr4 996015 996295 IDUA chr4 996512 996952 IDUA chr4 997080 997495 IDUA chr4 997750 997950 IDUA chr4 997993 998233 IDUA chr4 146560292 146560732 MMAA chr4 146563457 146563697 MMAA chr4 146567110 146567350 MMAA chr4 146572177 146572337 MMAA chr4 146575092 146575332 MMAA chr4 146576288 146576568 MMAA chr5 7870808 7871008 MTRR chr5 7873526 7873726 MTRR chr5 7875317 7875517 MTRR chr5 7877956 7878156 MTRR chr5 7878321 7878521 MTRR chr5 7883759 7883959 MTRR chr5 7889160 7889360 MTRR chr5 7891418 7891618 MTRR chr5 7892788 7892988 MTRR chr5 7895806 7896006 MTRR Chromosome Beginning End Condition chr5 7896923 7897437 MTRR chr5 35860853 35861185 IL7R chr5 35867359 35867599 IL7R chr5 35871124 35871364 IL7R chr5 35873525 35873805 IL7R chr5 35874493 35874653 IL7R chr5 35875989 35876189 IL7R chr5 70247697 70247977 SMN1 chr5 70888665 70888865 MCCC2 chr5 70892078 70892238 MCCC2 chr5 70895442 70895642 MCCC2 chr5 70898264 70898504 MCCC2 chr5 70900140 70900300 MCCC2 chr5 70922453 70922613 MCCC2 chr5 70930751 70930911 MCCC2 chr5 70930956 70931116 MCCC2 chr5 70936790 70936950 MCCC2 chr5 70939596 70939756 MCCC2 chr5 70942006 70942206 MCCC2 chr5 70944961 70945121 MCCC2 chr5 70945841 70946001 MCCC2 chr5 70948469 70948669 MCCC2 chr5 70952527 70952727 MCCC2 chr5 78076207 78076487 ARSB chr5 78077616 78077856 ARSB chr5 78135115 78135315 ARSB chr5 78181386 78181666 ARSB chr5 78251088 78251368 ARSB chr5 78260221 78260461 ARSB chr5 78264781 78265061 ARSB chr5 78280549 78280709 ARSB chr5 78280738 78280978 ARSB chr5 131705507 131706067 SLC22A5 chr5 131713829 131714218 SLC22A5 chr5 131719785 131720065 SLC22A5 chr5 131720965 131721245 SLC22A5 chr5 131722619 131722819 SLC22A5 chr5 131724563 131724763 SLC22A5 chr5 131726368 131726608 SLC22A5 chr5 131728112 131728352 SLC22A5 chr5 131729316 131729556 SLC22A5 chr5 131729783 131729983 SLC22A5 chr5 172659586 172659906 NKX2-5 chr5 172659911 172660231 NKX2-5 chr5 172661733 172662333 NKX2-5 chr5 177419687 177419887 PROP1 chr5 177419945 177420105 PROP1 chr5 177421104 177421344 PROP1 chr5 177422779 177422979 PROP1 chr6 32006219 32006459 CYP21A2 chr6 32006463 32006623 CYP21A2 chr6 32006807 32007007 CYP21A2 chr6 32007102 32007432 CYP21A2 chr6 32007489 32007689 CYP21A2 chr6 32007763 32008003 CYP21A2 chr6 32008100 32008300 CYP21A2 Chromosome Beginning End Condition chr6 32008308 32008548 CYP21A2 chr6 32008642 32008882 CYP21A2 chr6 49399453 49399613 MMUT chr6 49403139 49403379 MMUT chr6 49407858 49408098 MMUT chr6 49409535 49409735 MMUT chr6 49412317 49412477 MMUT chr6 49415341 49415541 MMUT chr6 49416476 49416716 MMUT chr6 49419164 49419444 MMUT chr6 49421273 49421553 MMUT chr6 49423772 49424012 MMUT chr6 49425383 49425783 MMUT chr6 49426785 49427225 MMUT chr6 80816336 80816616 BCKDHB chr6 80837162 80837362 BCKDHB chr6 80838847 80839007 BCKDHB chr6 80877348 80877508 BCKDHB chr6 80878550 80878790 BCKDHB chr6 80880955 80881155 BCKDHB chr6 80910603 80910803 BCKDHB chr6 80912772 80912932 BCKDHB chr6 80982795 80982995 BCKDHB chr6 81053354 81053554 BCKDHB chr6 150710553 150710713 IYD chr6 150715262 150715462 IYD chr7 65425904 65426104 GUSB chr7 65429324 65429484 GUSB chr7 65432701 65432941 GUSB chr7 65435216 65435416 GUSB chr7 65439253 65439453 GUSB chr7 65439491 65439731 GUSB chr7 65439862 65440062 GUSB chr7 65440935 65441135 GUSB chr7 65444364 65444564 GUSB chr7 65444711 65444951 GUSB chr7 65445238 65445438 GUSB chr7 65447015 65447215 GUSB chr7 65546731 65547011 ASL chr7 65547290 65547490 ASL chr7 65547814 65547974 ASL chr7 65548019 65548219 ASL chr7 65551570 65551810 ASL chr7 65552264 65552424 ASL chr7 65552652 65552852 ASL chr7 65553725 65553925 ASL chr7 65554043 65554203 ASL chr7 65554212 65554372 ASL chr7 65554543 65554743 ASL chr7 65556953 65557153 ASL chr7 65557501 65557701 ASL chr7 65557715 65557915 ASL chr7 107531621 107531821 DLD chr7 107533606 107533806 DLD chr7 107542154 107542314 DLD chr7 107542682 107542882 DLD Chromosome Beginning End Condition chr7 107543821 107544021 DLD chr7 107545305 107545505 DLD chr7 107546705 107546865 DLD chr7 107555907 107556187 DLD chr7 107557228 107557428 DLD chr7 107557710 107557950 DLD chr7 107558372 107558572 DLD chr7 107559403 107559774 DLD chr7 107571837 107572037 LAMB1 chr7 117120089 117120249 CFTR chr7 117144264 117144464 CFTR chr7 117149043 117149243 CFTR chr7 117170939 117171179 CFTR chr7 117174276 117174476 CFTR chr7 117175244 117175524 CFTR chr7 117176577 117176817 CFTR chr7 117178943 117179143 CFTR chr7 117180137 117180417 CFTR chr7 117182015 117182215 CFTR chr7 117188664 117188944 CFTR chr7 117199474 117199754 CFTR chr7 117227737 117227937 CFTR chr7 117229426 117229626 CFTR chr7 117230352 117230552 CFTR chr7 117231970 117232730 CFTR chr7 117234928 117235168 CFTR chr7 117242820 117242980 CFTR chr7 117243571 117243851 CFTR chr7 117246661 117246861 CFTR chr7 117250528 117250768 CFTR chr7 117251594 117252074 CFTR chr7 117252102 117252302 CFTR chr7 117254619 117254819 CFTR chr7 117267559 117267879 CFTR chr7 117279915 117280115 CFTR chr7 117282458 117283258 CFTR chr7 117292841 117293041 CFTR chr7 117304721 117305641 CFTR chr7 117306958 117307198 CFTR chr8 17914967 17915167 ASAH1 chr8 17916251 17916451 ASAH1 chr8 17916833 17916993 ASAH1 chr8 17917021 17917221 ASAH1 chr8 17918855 17919015 ASAH1 chr8 17919114 17919314 ASAH1 chr8 17919739 17919979 ASAH1 chr8 17920682 17920802 ASAH1 chr8 17921906 17922106 ASAH1 chr8 17924636 17924836 ASAH1 chr8 17927267 17927427 ASAH1 chr8 17928768 17928968 ASAH1 chr8 17932966 17933166 ASAH1 chr8 17941397 17941597 ASAH1 chr8 110099690 110099890 TRHR chr8 110099938 110100178 TRHR chr8 133880277 133880477 TG Chromosome Beginning End Condition chr8 133881961 133882161 TG chr8 133883490 133883690 TG chr8 133885207 133885567 TG chr8 133894754 133894954 TG chr8 133898646 133898806 TG chr8 133898859 133899059 TG chr8 133899135 133899335 TG chr8 133900353 133900553 TG chr8 133900659 133900859 TG chr8 133910954 133911154 TG chr8 133913605 133913805 TG chr8 133918981 133919141 TG chr8 133923576 133923776 TG chr8 133935542 133935742 TG chr8 133953638 133953838 TG chr8 133961073 133961273 TG chr8 133975185 133975385 TG chr8 133978843 133979043 TG chr8 133979942 133980255 TG chr8 133981634 133981834 TG chr8 133983950 133984150 TG chr8 133995494 133995694 TG chr8 134024162 134024362 TG chr8 134030085 134030285 TG chr8 134034273 134034473 TG chr8 134042052 134042252 TG chr8 134146834 134147034 TG chr8 143955726 143955926 CYP11B1 chr8 143956328 143956608 CYP11B1 chr8 143956616 143956776 CYP11B1 chr8 143957079 143957359 CYP11B1 chr8 143957599 143957839 CYP11B1 chr8 143958047 143958207 CYP11B1 chr8 143958331 143958722 CYP11B1 chr8 143960402 143960642 CYP11B1 chr8 143960959 143961239 CYP11B1 chr9 34646644 34646844 GALT chr9 34647030 34647310 GALT chr9 34647475 34647715 GALT chr9 34647774 34648014 GALT chr9 34648061 34648221 GALT chr9 34648272 34648512 GALT chr9 34648740 34649100 GALT chr9 34649391 34649631 GALT chr9 34650306 34650506 GALT chr9 100616308 100616600 FOXE1 chr9 104184036 104184236 ALDOB chr9 104187103 104187383 ALDOB chr9 104187763 104187963 ALDOB chr9 104188802 104188962 ALDOB chr9 104189728 104189968 ALDOB chr9 104190698 104190858 ALDOB chr9 104191955 104192296 ALDOB chr9 104192957 104193260 ALDOB chr9 104197890 104198090 ALDOB chr9 119460082 119460282 ASTN2 Chromosome Beginning End Condition chr9 119460309 119460776 ASTN2 chr9 119460838 119461038 ASTN2 chr9 119461075 119461275 ASTN2 chr9 119461369 119461609 ASTN2 chr9 119461649 119461849 ASTN2 chr9 133327564 133327724 ASS1 chr9 133329654 133329814 ASS1 chr9 133333738 133334018 ASS1 chr9 133339448 133339608 ASS1 chr9 133342062 133342222 ASS1 chr9 133346162 133346362 ASS1 chr9 133346774 133346974 ASS1 chr9 133352204 133352404 ASS1 chr9 133355049 133355289 ASS1 chr9 133355724 133355884 ASS1 chr9 133364668 133364908 ASS1 chr9 133370212 133370452 ASS1 chr9 133374836 133374996 ASS1 chr9 133376262 133376462 ASS1 chr9 139089393 139089593 LHX3 chr9 139090543 139090703 LHX3 chr9 139091415 139091750 LHX3 chr9 139092465 139092665 LHX3 chrlO 6061326 6061526 IL2RA chrlO 6063427 6063758 IL2RA chrlO 6066173 6066373 IL2RA chrlO 6067702 6068042 IL2RA chrlO 6103952 6104152 IL2RA chrlO 14950429 14951426 DCLRE1C chrlO 14968784 14968944 DCLRE1C chrlO 14969914 14970114 DCLRE1C chrlO 14974766 14974966 DCLRE1C chrlO 14976345 14976545 DCLRE1C chrlO 14976658 14976818 DCLRE1C chrlO 14977423 14977583 DCLRE1C chrlO 14978442 14978642 DCLRE1C chrlO 14987079 14987239 DCLRE1C chrlO 14995858 14996058 DCLRE1C chrlO 90974577 90974857 LIPA chrlO 90975641 90975801 LIPA chrlO 90982183 90982383 LIPA chrlO 90983354 90983684 LIPA chrlO 90984819 90984979 LIPA chrlO 90986610 90986810 LIPA chrlO 90987904 90988184 LIPA chrlO 91005421 91005581 LIPA chrlO 91007194 91007505 LIPA chrlO 91011392 91011592 LIPA chrlO 104590479 104590759 CYP17A1 chrlO 104591203 104591403 CYP17A1 chrlO 104592236 104592476 CYP17A1 chrlO 104592735 104592895 CYP17A1 chrlO 104593751 104593951 CYP17A1 chrlO 104594507 104594873 CYP17A1 chrlO 104594956 104595196 CYP17A1 chrlO 104596809 104597129 CYP17A1 Chromosome Beginning End Condition chrll 5246608 5247248 HBB chrll 5247791 5248791 HBB chrll 6411749 6411949 SMPD1 chrll 6411952 6412232 SMPD1 chrll 6412630 6413390 SMPD1 chrll 6414386 6414666 SMPD1 chrll 6414799 6414959 SMPD1 chrll 6415060 6415300 SMPD1 chrll 6415420 6415780 SMPD1 chrll 6635734 6635974 TPP1 chrll 6636055 6636255 TPP1 chrll 6636363 6636603 TPP1 chrll 6636614 6636854 TPP1 chrll 6637191 6637351 TPP1 chrll 6637538 6637738 TPP1 chrll 6637861 6638141 TPP1 chrll 6638155 6638435 TPP1 chrll 6638479 6638719 TPP1 chrll 6638810 6639050 TPP1 chrll 6639951 6640111 TPP1 chrll 6640322 6640714 TPP1 chrll 36595099 36595419 RAG1 chrll 36595515 36595715 RAG1 chrll 36595742 36595982 RAG1 chrll 36596024 36596544 RAG1 chrll 36596621 36596821 RAG1 chrll 36596851 36597211 RAG1 chrll 36597300 36597460 RAG1 chrll 36597524 36597844 RAG1 chrll 36614256 36614456 RAG2 chrll 36614715 36614915 RAG2 chrll 36615073 36615818 RAG2 chrll 71146415 71146895 DHCR7 chrll 71148803 71149043 DHCR7 chrll 71149907 71150147 DHCR7 chrll 71152260 71152500 DHCR7 chrll 71153253 71153453 DHCR7 chrll 71154981 71155141 DHCR7 chrll 71155155 71155315 DHCR7 chrll 71155847 71156047 DHCR7 chrll 107992294 107992454 AC ATI chrll 108002611 108002811 AC ATI chrll 108004489 108004689 AC ATI chrll 108004935 108005095 AC ATI chrll 108005829 108006029 AC ATI chrll 108009580 108009820 AC ATI chrll 108010789 108010989 AC ATI chrll 108012239 108012479 AC ATI chrll 108013100 108013300 AC ATI chrll 108014654 108014814 AC ATI chrll 108016867 108017147 AC ATI chrll 108017960 108018120 AC ATI chrll 118182786 118182986 CD3E chrll 118183303 118183543 CD3E chrll 118184475 118184675 CD3E chrll 118209830 118210030 CD3D Chromosome Beginning End Condition chrll 118210517 118210717 CD3D chrll 118211041 118211281 CD3D chrl2 103232895 103233055 PAH chrl2 103234117 103234357 PAH chrl2 103237348 103237628 PAH chrl2 103238060 103238260 PAH chrl2 103240623 103240783 PAH chrl2 103245419 103245579 PAH chrl2 103246544 103246784 PAH chrl2 103248196 103248436 PAH chrl2 103248453 103248573 PAH chrl2 103248605 103249125 PAH chrl2 103260330 103260490 PAH chrl2 103271182 103271382 PAH chrl2 103288470 103288750 PAH chrl2 103306516 103306676 PAH chrl2 103310796 103310956 PAH chrl2 103351946 103352146 ASCL1 chrl2 109994826 109994986 MMAB chrl2 109996851 109997011 MMAB chrl2 109998811 109998971 MMAB chrl2 109999177 109999337 MMAB chrl2 109999544 109999744 MMAB chrl2 110002880 110003040 MMAB chrl2 110006523 110006723 MMAB chrl2 110011132 110011332 MMAB chrl3 20763037 20763757 GJB2 chrl3 20766755 20766955 GJB2 chrl3 48877894 48878094 RBI chrl3 48878109 48878269 RBI chrl3 48881349 48881589 RBI chrl3 48916697 48916897 RBI chrl3 48919156 48919396 RBI chrl3 48921884 48922084 RBI chrl3 48923051 48923211 RBI chrl3 48934142 48934302 RBI chrl3 48936902 48937142 RBI chrl3 48938969 48939169 RBI chrl3 48941586 48941786 RBI chrl3 48942575 48942775 RBI chrl3 48947485 48947685 RBI chrl3 48951012 48951212 RBI chrl3 48953675 48953835 RBI chrl3 48954274 48954474 RBI chrl3 48955362 48955602 RBI chrl3 48985641 48985841 RBI chrl3 48985962 48986202 RBI chrl3 48986272 48986472 RBI chrl3 49027064 49027304 RBI chrl3 49030295 49030535 RBI chrl3 49033777 49034017 RBI chrl3 49037819 49038019 RBI chrl3 49039122 49039522 RBI chrl3 49045998 49046198 RBI chrl3 49047413 49047613 RBI chrl3 49050747 49051080 RBI Chromosome Beginning End Condition chrl3 49051419 49051619 RBI chrl3 52508895 52509175 ATP7B chrl3 52509681 52509881 ATP7B chrl3 52511394 52511834 ATP7B chrl3 52513145 52513345 ATP7B chrl3 52515163 52515403 ATP7B chrl3 52516472 52516752 ATP7B chrl3 52518230 52518990 ATP7B chrl3 52520382 52520662 ATP7B chrl3 52523746 52523986 ATP7B chrl3 52524101 52524341 ATP7B chrl3 52524356 52524556 ATP7B chrl3 52531597 52531797 ATP7B chrl3 52532428 52532708 ATP7B chrl3 52534239 52534519 ATP7B chrl3 52535904 52536104 ATP7B chrl3 52538972 52539252 ATP7B chrl3 52542515 52542795 ATP7B chrl3 52544611 52544851 ATP7B chrl3 52548051 52548851 ATP7B chrl3 52548951 52549311 ATP7B chrl3 52585365 52585525 ATP7B chrl3 52585804 52586004 ATP7B chrl3 100741328 100741528 PCCA chrl3 100755109 100755269 PCCA chrl3 100764039 100764353 PCCA chrl3 100807191 100807391 PCCA chrl3 100809486 100809646 PCCA chrl3 100861563 100861763 PCCA chrl3 100888010 100888210 PCCA chrl3 100909783 100909983 PCCA chrl3 100914971 100915091 PCCA chrl3 100920911 100921071 PCCA chrl3 100925405 100925645 PCCA chrl3 100953667 100953907 PCCA chrl3 100955145 100955345 PCCA chrl3 100959405 100959565 PCCA chrl3 100962051 100962251 PCCA chrl3 100982827 100983027 PCCA chrl3 100992383 100992583 PCCA chrl3 101020680 101020880 PCCA chrl3 101077835 101078035 PCCA chrl3 101101430 101101550 PCCA chrl3 101167651 101167891 PCCA chrl3 101179867 101180067 PCCA chrl3 101182290 101182450 PCCA chrl3 108860928 108861582 LIG4 chrl3 108861613 108861813 LIG4 chrl3 108861819 108862573 LIG4 chrl3 108862664 108862864 LIG4 chrl3 108862904 108863104 LIG4 chrl3 108863313 108863513 LIG4 chrl4 20940425 20940732 PNP chrl4 20942625 20943136 PNP chrl4 20943227 20943387 PNP chrl4 20944536 20944736 PNP Chromosome Beginning End Condition chrl4 23242787 23242947 SLC7A7 chrl4 23243112 23243352 SLC7A7 chrl4 23243533 23243733 SLC7A7 chrl4 23244599 23244799 SLC7A7 chrl4 23244995 23245195 SLC7A7 chrl4 23245360 23245560 SLC7A7 chrl4 23247959 23248119 SLC7A7 chrl4 23249074 23249274 SLC7A7 chrl4 23282098 23282618 SLC7A7 chrl4 36986383 36986732 NKX2-1 chrl4 36986748 36986868 NKX2-1 chrl4 36986886 36987246 NKX2-1 chrl4 36988174 36988414 NKX2-1 chrl4 36989234 36989434 NKX2-1 chrl4 81421995 81422195 TSHR chrl4 81534524 81534724 TSHR chrl4 81554206 81554406 TSHR chrl4 81557338 81557538 TSHR chrl4 81558843 81559003 TSHR chrl4 81562885 81563085 TSHR chrl4 81606023 81606223 TSHR chrl4 81609271 81609431 TSHR chrl4 81609555 81609995 TSHR chrl4 81610189 81610429 TSHR chrl4 88401038 88401278 GALC chrl4 88406185 88406385 GALC chrl4 88407711 88407951 GALC chrl4 88411842 88412082 GALC chrl4 88414019 88414219 GALC chrl4 88416143 88416303 GALC chrl4 88416968 88417168 GALC chrl4 88429673 88429873 GALC chrl4 88431796 88431996 GALC chrl4 88434619 88434819 GALC chrl4 88442646 88442886 GALC chrl4 88448486 88448646 GALC chrl4 88450691 88450931 GALC chrl4 88452788 88452988 GALC chrl4 88454387 88454587 GALC chrl4 88454753 88454913 GALC chrl4 88459287 88459527 GALC chrl4 88459675 88459875 GALC chrl5 40697977 40698217 IVD chrl5 40699782 40699982 IVD chrl5 40700046 40700246 IVD chrl5 40702810 40703570 IVD chrl5 40703703 40703903 IVD chrl5 40705170 40705330 IVD chrl5 40707035 40707235 IVD chrl5 40707563 40707723 IVD chrl5 40708220 40708380 IVD chrl5 40708444 40708604 IVD chrl5 40710305 40710505 IVD chrl5 45386339 45386539 DUOX2 chrl5 45389334 45389534 DUOX2 chrl5 45389744 45389944 DUOX2 Chromosome Beginning End Condition chrl5 45390157 45390357 DUOX2 chrl5 45391488 45391782 DUOX2 chrl5 45391880 45392080 DUOX2 chrl5 45392942 45393142 DUOX2 chrl5 45393323 45393523 DUOX2 chrl5 45394077 45394277 DUOX2 chrl5 45396059 45396259 DUOX2 chrl5 45396322 45396522 DUOX2 chrl5 45398315 45398625 DUOX2 chrl5 45398688 45398888 DUOX2 chrl5 45399548 45399748 DUOX2 chrl5 45400249 45400449 DUOX2 chrl5 45400985 45401185 DUOX2 chrl5 45402033 45402193 DUOX2 chrl5 45402588 45402788 DUOX2 chrl5 45403595 45403795 DUOX2 chrl5 45403910 45404190 DUOX2 chrl5 45405137 45405337 DUOX2 chrl5 45406711 45406911 DUOX2 chrl5 45408686 45408886 DUOXA2 chrl5 45409372 45409572 DUOXA2 chrl5 80445328 80445528 FAH chrl5 80450302 80450613 FAH chrl5 80452038 80452278 FAH chrl5 80452697 80452857 FAH chrl5 80454571 80454731 FAH chrl5 80460386 80460666 FAH chrl5 80464441 80464641 FAH chrl5 80465301 80465541 FAH chrl5 80467284 80467484 FAH chrl5 80469826 80470026 FAH chrl5 80472418 80472618 FAH chrl5 80473322 80473562 FAH chrl5 80478430 80478590 FAH chrl7 3543424 3543624 CTNS chrl7 3550677 3550877 CTNS chrl7 3552115 3552315 CTNS chrl7 3558257 3558457 CTNS chrl7 3558522 3558682 CTNS chrl7 3559782 3560102 CTNS chrl7 3561238 3561518 CTNS chrl7 3563109 3563309 CTNS chrl7 3563470 3563710 CTNS chrl7 7120393 7120593 DLG4 chrl7 7121019 7121179 DLG4 chrl7 7123292 7123532 ACADVL chrl7 7123770 7124010 ACADVL chrl7 7124090 7124370 ACADVL chrl7 7124799 7125039 ACADVL chrl7 7125229 7125429 ACADVL chrl7 7125453 7125653 ACADVL chrl7 7125947 7126227 ACADVL chrl7 7126414 7126614 ACADVL chrl7 7126929 7127409 ACADVL chrl7 7127451 7127611 ACADVL chrl7 7127644 7127884 ACADVL Chromosome Beginning End Condition chrl7 7127949 7128189 ACADVL chrl7 7128241 7128401 ACADVL chrl7 38233673 38233873 THRA chrl7 38242924 38243124 THRA chrl7 38244459 38244659 THRA chrl7 38245535 38245735 THRA chrl7 40688193 40688353 NAGLU chrl7 40688536 40688736 NAGLU chrl7 40689370 40689610 NAGLU chrl7 40690306 40690569 NAGLU chrl7 40690648 40690808 NAGLU chrl7 40692983 40693223 NAGLU chrl7 40695069 40695749 NAGLU chrl7 40695824 40696264 NAGLU chrl7 73748202 73748402 ITGB4 chrl7 73748404 73748604 ITGB4 chrl7 73750655 73750855 ITGB4 chrl7 73751788 73751988 ITGB4 chrl7 73753088 73753248 ITGB4 chrl7 73753250 73753617 ITGB4,GALK1 chrl7 73754072 73754469 GALK1 chrl7 73754475 73754675 GALK1 chrl7 73758770 73759010 GALK1 chrl7 73759366 73759566 GALK1 chrl7 73759985 73760145 GALK1 chrl7 73761055 73761295 GALK1 chrl7 78078336 78078936 GAA chrl7 78079514 78079754 GAA chrl7 78081265 78081769 GAA chrl7 78082112 78082632 GAA chrl7 78083702 78083902 GAA chrl7 78084507 78084827 GAA chrl7 78085724 78085964 GAA chrl7 78086335 78086535 GAA chrl7 78086634 78086874 GAA chrl7 78086975 78087215 GAA chrl7 78090729 78090969 GAA chrl7 78091382 78092622 GAA chrl7 78093024 78093184 GAA chrl7 78184255 78184815 SGSH chrl7 78185862 78186102 SGSH chrl7 78187570 78187730 SGSH chrl7 78187948 78188148 SGSH chrl7 78188378 78188618 SGSH chrl7 78188823 78188983 SGSH chrl7 78190799 78191039 SGSH chrl7 78194006 78194166 SGSH chrl9 1397301 1397501 GAMT chrl9 1398904 1399064 GAMT chrl9 1399090 1399250 GAMT chrl9 1399475 1399635 GAMT chrl9 1399746 1399986 GAMT chrl9 1401270 1401510 GAMT chrl9 12757372 12757572 MAN2B1 chrl9 12757997 12758157 MAN2B1 chrl9 12758215 12758455 MAN2B1 Chromosome Beginning End Condition chrl9 12758888 12759088 MAN2B1 chrl9 12759118 12759318 MAN2B1 chrl9 12759890 12760090 MAN2B1 chrl9 12760099 12760299 MAN2B1 chrl9 12760731 12761051 MAN2B1 chrl9 12762961 12763281 MAN2B1 chrl9 12766461 12766741 MAN2B1 chrl9 12767394 12767554 MAN2B1 chrl9 12767712 12767872 MAN2B1 chrl9 12768223 12768423 MAN2B1 chrl9 12768791 12768991 MAN2B1 chrl9 12769054 12769294 MAN2B1 chrl9 12771972 12772172 MAN2B1 chrl9 12774165 12774365 MAN2B1 chrl9 12774452 12774652 MAN2B1 chrl9 12775556 12775890 MAN2B1 chrl9 12776117 12776425 MAN2B1 chrl9 12776453 12776653 MAN2B1 chrl9 12777295 12777575 MAN2B1 chrl9 13002067 13002267 GCDH chrl9 13002608 13002848 GCDH chrl9 13002880 13003040 GCDH chrl9 13004247 13004527 GCDH chrl9 13006763 13006923 GCDH chrl9 13007003 13007243 GCDH chrl9 13007677 13007877 GCDH chrl9 13008100 13008300 GCDH chrl9 13008481 13008721 GCDH chrl9 13010238 13010398 GCDH chrl9 17937608 17937808 JAK3 chrl9 17940922 17941122 JAK3 chrl9 17941319 17941519 JAK3 chrl9 17941997 17942197 JAK3 chrl9 17942495 17942695 JAK3 chrl9 17945307 17945507 JAK3 chrl9 17945617 17945817 JAK3 chrl9 17945870 17946070 JAK3 chrl9 17946691 17946851 JAK3 chrl9 17947903 17948063 JAK3 chrl9 17948692 17948852 JAK3 chrl9 17949028 17949188 JAK3 chrl9 17950245 17950445 JAK3 chrl9 17950993 17951193 JAK3 chrl9 17952467 17952627 JAK3 chrl9 17953208 17953408 JAK3 chrl9 17953826 17954026 JAK3 chrl9 17954160 17954360 JAK3 chrl9 17954531 17954731 JAK3 chrl9 17955003 17955163 JAK3 chrl9 17983305 17983505 SLC5A5 chrl9 17988541 17988741 SLC5A5 chrl9 17992670 17993069 SLC5A5 chrl9 17999124 17999324 SLC5A5 chrl9 41903690 41903890 BCKDHA chrl9 41916459 41916990 BCKDHA chrl9 41919920 41920120 BCKDHA Chromosome Beginning End Condition chrl9 41925005 41925245 BCKDHA chrl9 41928052 41928292 BCKDHA chrl9 41928482 41928722 BCKDHA chrl9 41928852 41929132 BCKDHA chrl9 41930295 41930535 BCKDHA chr20 43248403 43248603 ADA chr20 43248887 43249127 ADA chr20 43249613 43249813 ADA chr20 43251181 43251341 ADA chr20 43251462 43251702 ADA chr20 43252790 43253030 ADA chr20 43254165 43254365 ADA chr20 43255046 43255286 ADA chr20 43257587 43257787 ADA chr20 43264807 43264967 ADA chr20 43280132 43280332 ADA chr21 38126497 38126777 HLCS chr21 38128800 38129080 HLCS chr21 38132025 38132185 HLCS chr21 38137272 38137512 HLCS chr21 38139420 38139620 HLCS chr21 38269102 38269493 HLCS chr21 38302493 38302733 HLCS chr21 38308668 38308868 HLCS chr21 38308922 38309202 HLCS chr21 38309229 38309715 HLCS chr21 38311084 38311284 HLCS chr21 44473913 44474153 CBS chr21 44476855 44477055 CBS chr21 44478264 44478464 CBS chr21 44478891 44479131 CBS chr21 44479270 44479470 CBS chr21 44480505 44480705 CBS chr21 44482361 44482561 CBS chr21 44483011 44483291 CBS chr21 44483956 44484156 CBS chr21 44485254 44485454 CBS chr21 44485489 44485969 CBS chr21 44486296 44486536 CBS chr21 44488560 44488760 CBS chr21 44492046 44492385 CBS chr21 44495774 44495974 CBS chr22 31006780 31007138 TCN2 chr22 31008929 31009129 TCN2 chr22 31010235 31010570 TCN2 chr22 31011012 31011212 TCN2 chr22 31011289 31011449 TCN2 chr22 31011501 31011813 TCN2 chr22 31013375 31013575 TCN2 chr22 31018876 31019076 TCN2 chr22 51063606 51063886 ARSA chr22 51063959 51064159 ARSA chr22 51064355 51064715 ARSA chr22 51065010 51065490 ARSA chr22 51065574 51065854 ARSA chr22 51065973 51066253 ARSA Chromosome Beginning End Condition chrX 30322666 30323066 NR0B1 chrX 30326278 30327398 NR0B1 chrX 31144664 31144864 DMD chrX 31165373 31165653 DMD chrX 31187523 31187763 DMD chrX 31190412 31190612 DMD chrX 31191593 31191793 DMD chrX 31196025 31196265 DMD chrX 31196735 31196975 DMD chrX 31198443 31198643 DMD chrX 31200580 31200780 DMD chrX 31200801 31201081 DMD chrX 31222016 31222296 DMD chrX 31224640 31224840 DMD chrX 31227571 31227851 DMD chrX 31241112 31241312 DMD chrX 31279008 31279208 DMD chrX 31279318 31279518 DMD chrX 31279680 31279880 DMD chrX 31341665 31341825 DMD chrX 31366612 31366812 DMD chrX 31462590 31462790 DMD chrX 31496218 31496498 DMD chrX 31497067 31497267 DMD chrX 31514864 31515104 DMD chrX 31525344 31525624 DMD chrX 31526227 31526427 DMD chrX 31627638 31627838 DMD chrX 31645741 31646021 DMD chrX 31676053 31676293 DMD chrX 31697478 31697718 DMD chrX 31747696 31747896 DMD chrX 31792118 31792358 DMD chrX 31838046 31838206 DMD chrX 31854794 31854994 DMD chrX 31893263 31893543 DMD chrX 31947654 31947894 DMD chrX 31950151 31950391 DMD chrX 31983046 31983246 DMD chrX 31986406 31986646 DMD chrX 32234993 32235233 DMD chrX 32305592 32305872 DMD chrX 32328191 32328431 DMD chrX 32360157 32360437 DMD chrX 32361203 32361443 DMD chrX 32364005 32364245 DMD chrX 32366480 32366680 DMD chrX 32380870 32381110 DMD chrX 32382643 32382883 DMD chrX 32383088 32383368 DMD chrX 32398572 32398852 DMD chrX 32404390 32404630 DMD chrX 32407559 32407839 DMD chrX 32408149 32408349 DMD chrX 32429811 32430091 DMD chrX 32456314 32456554 DMD Chromosome Beginning End Condition chrX 32459239 32459479 DMD chrX 32466525 32466805 DMD chrX 32472757 32473037 DMD chrX 32479420 32479620 DMD chrX 32481493 32481773 DMD chrX 32482646 32482846 DMD chrX 32486604 32486844 DMD chrX 32490234 32490474 DMD chrX 32502985 32503265 DMD chrX 32509375 32509655 DMD chrX 32519814 32520014 DMD chrX 32536061 32536301 DMD chrX 32563225 32563505 DMD chrX 32582844 32583439 DMD chrX 32583445 32583965 DMD chrX 32591601 32591801 DMD chrX 32591813 32592013 DMD chrX 32613814 32614054 DMD chrX 32632359 32632639 DMD chrX 32662182 32662532 DMD chrX 32663035 32663315 DMD chrX 32715980 32716180 DMD chrX 32717180 32717460 DMD chrX 32756808 32757008 DMD chrX 32827548 32827788 DMD chrX 32834536 32834816 DMD chrX 32841359 32841559 DMD chrX 32841867 32842067 DMD chrX 32862837 32863037 DMD chrX 32867791 32867991 DMD chrX 33038199 33038359 DMD chrX 33192352 33192552 DMD chrX 33229308 33229508 DMD chrX 70327626 70327826 IL2RG chrX 70328082 70328242 IL2RG chrX 70328398 70328598 IL2RG chrX 70329053 70329293 IL2RG chrX 70329954 70330194 IL2RG chrX 70330333 70330573 IL2RG chrX 70330698 70330898 IL2RG chrX 70331239 70331439 IL2RG chrX 77227068 77227268 ATP7A chrX 77243633 77243833 ATP7A chrX 77243968 77244317 ATP7A chrX 77244894 77245399 ATP7A chrX 77253944 77254224 ATP7A chrX 77258513 77258793 ATP7A chrX 77264618 77264778 ATP7A chrX 77266613 77266813 ATP7A chrX 77266895 77267207 ATP7A chrX 77268372 77268612 ATP7A chrX 77270113 77270313 ATP7A chrX 77271193 77271433 ATP7A chrX 77275698 77275938 ATP7A chrX 77276438 77276638 ATP7A chrX 77284715 77284995 ATP7A Chromosome Beginning End Condition chrX 77286813 77287139 ATP7A chrX 77289089 77289369 ATP7A chrX 77294257 77294457 ATP7A chrX 77296134 77296294 ATP7A chrX 77298065 77298305 ATP7A chrX 77298774 77298974 ATP7A chrX 77300924 77301124 ATP7A chrX 77301814 77302014 ATP7A chrX 100652789 100653109 GLA chrX 100653339 100653579 GLA chrX 100653713 100653993 GLA chrX 100655601 100655801 GLA chrX 100656570 100656850 GLA chrX 100658746 100659026 GLA chrX 100662654 100662934 GLA chrX 148564325 148564765 IDS chrX 148568443 148568683 IDS chrX 148568744 148568944 IDS chrX 148571786 148572026 IDS chrX 148577820 148578060 IDS chrX 148579618 148579858 IDS chrX 148582457 148582617 IDS chrX 148584793 148585073 IDS chrX 148585642 148585842 IDS chrX 148586544 148586904 IDS chrX 152990703 152991623 ABCD1 chrX 152994647 152994927 ABCD1 chrX 153001565 153001965 ABCD1 chrX 153002562 153002722 ABCD1 chrX 153005499 153005739 ABCD1 chrX 153005981 153006221 ABCD1 chrX 153008383 153008583 ABCD1 chrX 153008613 153008853 ABCD1 chrX 153008886 153009272 ABCD1 chrX 153760198 153760722 G6PD chrX 153760786 153761066 G6PD chrX 153761151 153761391 G6PD chrX 153761743 153761903 G6PD chrX 153762257 153762457 G6PD chrX 153762515 153762755 G6PD chrX 153763373 153763533 G6PD chrX 153764094 153764294 G6PD chrX 153774163 153774363 G6PD

Table 3. Target regions for the PD5 179 Gene Newborn Screening Panel

Chromosome Region Region Condition chrl 11850728 11850968 MTHFR chrl 11851214 11851382 MTHFR chrl 11852273 11852489 MTHFR chrl 11853914 11854178 MTHFR chrl 11854388 11854652 MTHFR chrl 11854734 11854974 MTHFR chrl 11855123 11855431 MTHFR chrl 11856203 11856515 MTHFR chrl 11860200 11860416 MTHFR chrl 11861221 11861461 MTHFR chrl 11862932 11863220 MTHFR chrl 19200862 19201078 ALDH4A1 chrl 19202804 19203020 ALDH4A1 chrl 19203884 19204100 ALDH4A1 chrl 19208085 19208301 ALDH4A1 chrl 19212015 19212231 ALDH4A1 chrl 19228889 19229105 ALDH4A1 chrl 21880543 21880687 ALPL chrl 21887090 21887282 ALPL chrl 21887540 21887756 ALPL chrl 21889547 21889787 ALPL chrl 21890496 21890760 ALPL chrl 21894549 21894789 ALPL chrl 21896757 21896925 ALPL chrl 21900107 21900347 ALPL chrl 21902174 21902462 ALPL chrl 21902968 21903184 ALPL chrl 21903879 21904143 ALPL chrl 24122636 24122780 GALE chrl 24122971 24123322 GALE chrl 24123510 24123726 GALE chrl 24124149 24124365 GALE chrl 24124598 24124742 GALE chrl 24125079 24125247 GALE chrl 24125290 24125506 GALE chrl 24128923 24129067 HMGCL chrl 24130849 24131017 HMGCL chrl 24134626 24134866 HMGCL chrl 24137202 24137346 HMGCL chrl 24140628 24140820 HMGCL chrl 24143119 24143335 HMGCL chrl 24143895 24144111 HMGCL chrl 24146946 24147090 HMGCL chrl 24151790 24151958 HMGCL chrl 32745221 32745793 LCK chrl 43392729 43392969 SLC2A1 chrl 43393224 43393536 SLC2A1 chrl 43394583 43394751 SLC2A1 chrl 43394797 43395037 SLC2A1 chrl 43395259 43395715 SLC2A1 chrl 43396299 43396539 SLC2A1 chrl 43396667 43396931 SLC2A1 chrl 43408852 43409044 SLC2A1 chrl 43424212 43424428 SLC2A1 chrl 45965950 45966142 MMACHC chrl 45972969 45973281 MMACHC chrl 45973832 45974096 MMACHC chrl 45974458 45974890 MMACHC chrl 45976980 45977196 MMACHC chrl 53662599 53662815 CPT2 chrl 53666350 53666494 CPT2 chrl 53667945 53668113 CPT2 chrl 53675681 53677265 CPT2 chrl 53678953 53679265 CPT2 chrl 76190414 76190558 ACADM chrl 76194042 76194234 ACADM chrl 76198270 76198669 ACADM chrl 76199155 76199371 ACADM chrl 76200424 76200616 ACADM chrl 76205610 76205850 ACADM chrl 76211407 76211647 ACADM chrl 76215054 76215294 ACADM chrl 76216083 76216275 ACADM chrl 76226797 76227061 ACADM chrl 76228323 76228491 ACADM chrl 100661718 100662006 DBT chrl 100671661 100671877 DBT chrl 100671991 100672231 DBT chrl 100676141 100676357 DBT chrl 100680314 100680602 DBT chrl 100681523 100681787 DBT chrl 100684073 100684353 DBT chrl 100696232 100696520 DBT chrl 100700944 100701160 DBT chrl 100706300 100706468 DBT chrl 100715217 100715433 DBT chrl 115576018 115576210 TSHB chrl 115576528 115576912 TSHB chrl 119957966 119958182 HSD3B2 chrl 119962086 119962254 HSD3B2 chrl 119964498 119965002 HSD3B2 chrl 119965077 119965293 HSD3B2 chrl 155204788 155205100 GBA chrl 155205421 155205685 GBA chrl 155205982 155206294 GBA chrl 155207139 155207379 GBA chrl 155207881 155208145 GBA chrl 155208256 155208496 GBA chrl 155209373 155209565 GBA chrl 155209666 155209882 GBA chrl 155210370 155210514 GBA chrl 155260294 155260510 PKLR chrl 155261521 155261761 PKLR chrl 155262966 155263350 PKLR chrl 155263988 155264204 PKLR chrl 155264279 155264519 PKLR chrl 155264912 155265128 PKLR chrl 155265179 155265395 PKLR chrl 155265432 155265576 PKLR chrl 155269781 155270122 PKLR chrl 198671552 198671768 PTPRC chrl 198685862 198686078 PTPRC chrl 198687288 198687504 PTPRC chrl 198704246 198704414 PTPRC chrl 236958897 236959113 MTR chrl 236969335 236969551 MTR chrl 236971941 236972157 MTR chrl 236987412 236987628 MTR chrl 236988592 236988808 MTR chrl 236998778 236998994 MTR chrl 237013589 237013805 MTR chrl 237015770 237015986 MTR chrl 237023069 237023285 MTR chrl 237024403 237024619 MTR chrl 237026645 237026861 MTR chrl 237038014 237038230 MTR chrl 237044041 237044185 MTR chrl 237052492 237052708 MTR chrl 237057723 237057939 MTR chrl 237058635 237058827 MTR chrl 237060206 237060422 MTR chr2 1439975 1440119 TPO chr2 1459885 1460053 TPO chr2 1481114 1481330 TPO chr2 1488269 1488664 TPO chr2 1491505 1491871 TPO chr2 1497501 1497858 TPO chr2 1499723 1499939 TPO chr2 1500369 1500513 TPO chr2 1507678 1507894 TPO chr2 26414079 26414247 HADHA chr2 26414291 26414531 HADHA chr2 26415125 26415317 HADHA chr2 26416396 26416708 HADHA chr2 26417380 26417548 HADHA chr2 26417896 26418112 HADHA chr2 26420502 26420670 HADHA chr2 26423958 26424290 HADHA chr2 26426901 26427117 HADHA chr2 26432624 26432768 HADHA chr2 26435388 26435556 HADHA chr2 26437299 26437491 HADHA chr2 26437960 26438104 HADHA chr2 26453003 26453219 HADHA chr2 26455002 26455146 HADHA chr2 26457034 26457274 HADHA chr2 26459706 26459850 HADHA chr2 26461741 26462115 HADHA chr2 26467349 26467517 HADHA chr2 26476994 26477210 HADHB chr2 26486262 26486406 HADHB chr2 26492771 26492915 HADHB chr2 26496409 26496625 HADHB chr2 26499835 26500051 HADHB chr2 26501493 26501709 HADHB chr2 26501949 26502373 HADHB chr2 26502834 26503050 HADHB chr2 26505808 26506024 HADHB chr2 26507722 26507866 HADHB chr2 26508259 26508475 HADHB chr2 26512676 26512892 HADHB chr2 71351467 71351683 MCEE chr2 73114482 73114698 SPR chr2 73114757 73114973 SPR chr2 73115432 73115672 SPR chr2 73118425 73118713 SPR chr2 84652594 84652810 SUCLG1 chr2 84658572 84658740 SUCLG1 chr2 84668104 84668320 SUCLG1 chr2 84668340 84668508 SUCLG1 chr2 84670372 84670540 SUCLG1 chr2 84676713 84676881 SUCLG1 chr2 84686240 84686408 SUCLG1 chr2 98340394 98340832 ZAP70 chr2 98349248 98349464 ZAP70 chr2 98349651 98350150 ZAP70 chr2 98351064 98351280 ZAP70 chr2 98351822 98351966 ZAP70 chr2 98353932 98354148 ZAP70 chr2 98354197 98354341 ZAP70 chr2 98354390 98354606 ZAP70 chr2 98355741 98355957 ZAP70 chr2 113984688 113984880 PAX8 chr2 113992950 113993166 PAX8 chr2 113999139 113999355 PAX8 chr2 114001963 114002179 PAX8 chr2 114004288 114004480 PAX8 chr2 150426546 150426690 MMADHC chr2 150427487 150427703 MMADHC chr2 150432176 150432392 MMADHC chr2 150432900 150433068 MMADHC chr2 150435943 150436207 MMADHC chr2 150438581 150438845 MMADHC chr2 189839137 189839353 C0L3A1 chr2 189849434 189849797 C0L3A1 chr2 189850339 189850555 C0L3A1 chr2 189851759 189851927 C0L3A1 chr2 189852757 189852925 C0L3A1 chr2 189853262 189853430 C0L3A1 chr2 189854066 189854234 C0L3A1 chr2 189854760 189854928 C0L3A1 chr2 189854977 189855145 C0L3A1 chr2 189855674 189855842 C0L3A1 chr2 189856150 189856509 C0L3A1 chr2 189856862 189857006 C0L3A1 chr2 189857554 189857698 C0L3A1 chr2 189858031 189858247 C0L3A1 chr2 189858712 189858880 C0L3A1 chr2 189858910 189859102 C0L3A1 chr2 189859211 189859379 C0L3A1 chr2 189859408 189859576 C0L3A1 chr2 189859709 189859877 C0L3A1 chr2 189860384 189860528 C0L3A1 chr2 189860794 189860962 C0L3A1 chr2 189861085 189861277 C0L3A1 chr2 189861829 189861997 C0L3A1 chr2 189862007 189862175 C0L3A1 chr2 189862369 189862537 C0L3A1 chr2 189862938 189863106 C0L3A1 chr2 189863335 189863503 C0L3A1 chr2 189864001 189864313 C0L3A1 chr2 189864512 189864680 C0L3A1 chr2 189866096 189866360 C0L3A1 chr2 189866966 189867134 C0L3A1 chr2 189867634 189867826 C0L3A1 chr2 189868082 189868250 C0L3A1 chr2 189868405 189868573 COL3A1 chr2 189868665 189869145 COL3A1 chr2 189870025 189870241 COL3A1 chr2 189870932 189871172 COL3A1 chr2 189871571 189871739 COL3A1 chr2 189872169 189872385 COL3A1 chr2 189872603 189872867 COL3A1 chr2 189873594 189873762 COL3A1 chr2 189873840 189874056 COL3A1 chr2 189874856 189875141 COL3A1 chr2 189875362 189875626 COL3A1 chr2 189876312 189876552 COL3A1 chr2 211342381 211342597 CPS1 chr2 211421363 211421692 CPS1 chr2 211437971 211438187 CPS1 chr2 211441025 211441265 CPS1 chr2 211444403 211444547 CPS1 chr2 211447319 211447463 CPS1 chr2 211452764 211452980 CPS1 chr2 211454774 211455014 CPS1 chr2 211456556 211456748 CPS1 chr2 211457505 211457721 CPS1 chr2 211459199 211459391 CPS1 chr2 211460186 211460354 CPS1 chr2 211464040 211464372 CPS1 chr2 211465240 211465456 CPS1 chr2 211466918 211467110 CPS1 chr2 211469824 211469992 CPS1 chr2 211471396 211471708 CPS1 chr2 211473029 211473173 CPS1 chr2 211473174 211473342 CPS1 chr2 211476787 211477051 CPS1 chr2 211481066 211481282 CPS1 chr2 211502413 211502605 CPS1 chr2 211503830 211503998 CPS1 chr2 211504699 211504843 CPS1 chr2 211507122 211507410 CPS1 chr2 211512574 211512766 CPS1 chr2 211513124 211513340 CPS1 chr2 211515038 211515254 CPS1 chr2 211518695 211518887 CPS1 chr2 211521191 211521359 CPS1 chr2 211523291 211523435 CPS1 chr2 211525119 211525469 CPS1 chr2 211527839 211527983 CPS1 chr2 211532882 211533098 CPS1 chr2 211539528 211539744 CPS1 chr2 211540458 211540674 CPS1 chr2 211541706 211541922 CPS1 chr2 219646857 219647206 CYP27A1 chr2 219674239 219674551 CYP27A1 chr2 219676916 219677204 CYP27A1 chr2 219677218 219677530 CYP27A1 chr2 219677588 219677876 CYP27A1 chr2 219678680 219679014 CYP27A1 chr2 219679091 219679451 CYP27A1 chr2 219679582 219679798 CYP27A1 chr2 219942818 219942986 NHEJ1 chr2 220011368 220011512 NHEJ1 chr2 220022143 220022311 NHEJ1 chr2 220022803 220023181 NHEJ1 chr2 241808233 241808473 AG XT chr2 241808528 241808840 AG XT chr2 241810008 241810176 AG XT chr2 241810708 241810924 AG XT chr2 241812347 241812515 AG XT chr2 241813343 241813535 AG XT chr2 241813953 241814697 AG XT chr2 241815290 241815482 AG XT chr2 241816893 241817109 AG XT chr2 241817383 241817623 AG XT chr2 241818077 241818269 AG XT chr3 15643309 15643453 HACL1 chr3 15676934 15677246 BTD chr3 15683357 15683621 BTD chr3 15685819 15686995 BTD chr3 33038505 33038649 GLB1 chr3 33038715 33038883 GLB1 chr3 33055545 33055809 GLB1 chr3 33058148 33058364 GLB1 chr3 33059887 33060103 GLB1 chr3 33063007 33063199 GLB1 chr3 33065700 33065868 GLB1 chr3 33087557 33087749 GLB1 chr3 33093241 33093505 GLB1 chr3 33094918 33095062 GLB1 chr3 33099526 33099814 GLB1 chr3 33106902 33107094 GLB1 chr3 33109668 33109836 GLB1 chr3 33110254 33110518 GLB1 chr3 33113980 33114244 GLB1 chr3 33118551 33118767 GLB1 chr3 33138443 33138635 GLB1 chr3 48895043 48895259 SLC25A20 chr3 48895861 48896077 SLC25A20 chr3 48896433 48896649 SLC25A20 chr3 48896903 48897119 SLC25A20 chr3 48899924 48900068 SLC25A20 chr3 48921378 48921618 SLC25A20 chr3 48929399 48929615 SLC25A20 chr3 48936090 48936282 SLC25A20 chr3 49454928 49455096 AMT chr3 49455147 49455411 AMT chr3 49456357 49456597 AMT chr3 49456624 49456864 AMT chr3 49457103 49457271 AMT chr3 49457585 49457825 AMT chr3 49458870 49459062 AMT chr3 49459485 49459725 AMT chr3 49459772 49459988 AMT chr3 87309093 87309285 P0U1F1 chr3 87310369 87310537 P0U1F1 chr3 87311192 87311408 POU1F1 chr3 87313387 87313531 POU1F1 chr3 87313557 87313773 POU1F1 chr3 87322524 87322740 POU1F1 chr3 87325434 87325650 POU1F1 chr3 135969117 135969461 PCCB chr3 135974590 135974854 PCCB chr3 135975354 135975522 PCCB chr3 135979284 135979428 PCCB chr3 135980767 135980935 PCCB chr3 136002626 136002842 PCCB chr3 136003081 136003297 PCCB chr3 136012571 136012763 PCCB chr3 136016734 136016974 PCCB chr3 136019816 136020008 PCCB chr3 136035683 136036016 PCCB chr3 136045584 136045776 PCCB chr3 136045956 136046148 PCCB chr3 136046417 136046633 PCCB chr3 136047582 136047750 PCCB chr3 136048694 136048910 PCCB chr3 180702371 180702587 DNAJC19 chr3 180704703 180704919 DNAJC19 chr3 180705910 180706126 DNAJC19 chr3 180707282 180707498 DNAJC19 chr3 182733231 182733375 MCCC1 chr3 182737832 182738048 MCCC1 chr3 182740185 182740401 MCCC1 chr3 182743487 182743703 MCCC1 chr3 182751728 182751920 MCCC1 chr3 182754946 182755328 MCCC1 chr3 182756768 182756984 MCCC1 chr3 182759302 182759566 MCCC1 chr3 182763152 182763368 MCCC1 chr3 182769849 182770065 MCCC1 chr3 182775054 182775198 MCCC1 chr3 182788799 182789087 MCCC1 chr3 182804422 182804566 MCCC1 chr3 182810216 182810384 MCCC1 chr3 182812286 182812502 MCCC1 chr3 182817088 182817280 MCCC1 chr4 980776 981123 IDUA chr4 981524 981788 IDUA chr4 983546 983762 IDUA chr4 984830 985046 IDUA chr4 994347 994539 IDUA chr4 994615 994831 IDUA chr4 995196 996004 IDUA chr4 996011 996299 IDUA chr4 996516 996948 IDUA chr4 997080 997320 IDUA chr4 997323 997467 IDUA chr4 997742 997958 IDUA chr4 997993 998233 IDUA chr4 17488720 17488936 QDPR chr4 17492262 17492478 QDPR chr4 17493839 17494007 QDPR chr4 17503299 17503515 QDPR chr4 17505919 17506135 QDPR chr4 17510868 17511036 QDPR chr4 17513542 17513734 QDPR chr4 108911098 108911314 HADH chr4 108930903 108931095 HADH chr4 108935616 108935760 HADH chr4 108944562 108944778 HADH chr4 108945082 108945298 HADH chr4 108948805 108949021 HADH chr4 108954287 108954503 HADH chr4 146560284 146560740 MMAA chr4 146563457 146563697 MMAA chr4 146567098 146567362 MMAA chr4 146572173 146572341 MMAA chr4 146575092 146575332 MMAA chr4 146576296 146576560 MMAA chr4 159593554 159593698 ETFDH chr4 159601578 159601818 ETFDH chr4 159603348 159603588 ETFDH chr4 159605699 159605891 ETFDH chr4 159606232 159606400 ETFDH chr4 159611443 159611659 ETFDH chr4 159616626 159616842 ETFDH chr4 159618685 159618901 ETFDH chr4 159620079 159620343 ETFDH chr4 159624515 159624803 ETFDH chr4 159627323 159627563 ETFDH chr4 159627772 159628012 ETFDH chr4 159629544 159629712 ETFDH chr5 7870800 7871016 MTRR chr5 7873518 7873734 MTRR chr5 7875309 7875525 MTRR chr5 7877948 7878164 MTRR chr5 7878313 7878529 MTRR chr5 7883751 7883967 MTRR chr5 7889188 7889332 MTRR chr5 7891410 7891626 MTRR chr5 7892792 7892984 MTRR chr5 7895810 7896002 MTRR chr5 7896927 7897445 MTRR chr5 35860845 35861193 IL7R chr5 35867359 35867599 IL7R chr5 35871112 35871376 IL7R chr5 35873533 35873797 IL7R chr5 35874501 35874645 IL7R chr5 35875981 35876197 IL7R chr5 36200216 36200432 NADK2 chr5 36207158 36207374 NADK2 chr5 36241833 36242049 NADK2 chr5 70247693 70247981 SMN1 chr5 70888657 70888873 MCCC2 chr5 70892074 70892242 MCCC2 chr5 70895446 70895638 MCCC2 chr5 70898264 70898504 MCCC2 chr5 70900136 70900304 MCCC2 chr5 70922437 70922629 MCCC2 chr5 70930747 70930915 MCCC2 chr5 70930952 70931120 MCCC2 chr5 70936786 70936954 MCCC2 chr5 70939604 70939748 MCCC2 chr5 70941998 70942214 MCCC2 chr5 70944957 70945125 MCCC2 chr5 70945837 70946005 MCCC2 chr5 70948497 70948641 MCCC2 chr5 70952519 70952735 MCCC2 chr5 78076215 78076479 ARSB chr5 78077616 78077856 ARSB chr5 78135119 78135311 ARSB chr5 78181394 78181658 ARSB chr5 78251072 78251384 ARSB chr5 78260209 78260473 ARSB chr5 78264777 78265065 ARSB chr5 78280560 78280704 ARSB chr5 78280732 78281044 ARSB chr5 118788217 118788385 HSD17B4 chr5 118788400 118788568 HSD17B4 chr5 118791910 118792126 HSD17B4 chr5 118809549 118809765 HSD17B4 chr5 118810047 118810191 HSD17B4 chr5 118811341 118811622 HSD17B4 chr5 118813085 118813253 HSD17B4 chr5 118814470 118814758 HSD17B4 chr5 118824835 118825027 HSD17B4 chr5 118829458 118829698 HSD17B4 chr5 118832189 118832357 HSD17B4 chr5 118835009 118835249 HSD17B4 chr5 118837677 118837845 HSD17B4 chr5 118842465 118842609 HSD17B4 chr5 118844781 118844925 HSD17B4 chr5 118850569 118850785 HSD17B4 chr5 118860860 118861004 HSD17B4 chr5 118861557 118861725 HSD17B4 chr5 118862785 118862953 HSD17B4 chr5 118866961 118867153 HSD17B4 chr5 118872061 118872301 HSD17B4 chr5 118877546 118877714 HSD17B4 chr5 131705511 131706063 SLC22A5 chr5 131713821 131714226 SLC22A5 chr5 131719781 131720069 SLC22A5 chr5 131720961 131721249 SLC22A5 chr5 131722611 131722827 SLC22A5 chr5 131724555 131724771 SLC22A5 chr5 131726368 131726608 SLC22A5 chr5 131728100 131728364 SLC22A5 chr5 131729316 131729556 SLC22A5 chr5 131729775 131729991 SLC22A5 chr5 172659578 172659914 NKX2-5 chr5 172659915 172660227 NKX2-5 chr5 172661745 172662321 NKX2-5 chr5 177419691 177419883 PR0P1 chr5 177419941 177420109 PR0P1 chr5 177421104 177421344 PROP1 chr5 177422771 177422987 PROP1 chr6 32006219 32006623 CYP21A2 chr6 32006799 32007015 CYP21A2 chr6 32007094 32007440 CYP21A2 chr6 32007481 32007697 CYP21A2 chr6 32007727 32008039 CYP21A2 chr6 32008092 32008548 CYP21A2 chr6 32008642 32008882 CYP21A2 chr6 42928546 42928762 GNMT chr6 42930479 42930695 GNMT chr6 42930767 42930983 GNMT chr6 49399437 49399629 MMUT chr6 49403127 49403391 MMUT chr6 49407858 49408098 MMUT chr6 49409527 49409743 MMUT chr6 49412313 49412481 MMUT chr6 49415333 49415549 MMUT chr6 49416476 49416716 MMUT chr6 49419172 49419436 MMUT chr6 49421273 49421561 MMUT chr6 49423772 49424012 MMUT chr6 49425391 49425775 MMUT chr6 49426789 49427221 MMUT chr6 70410581 70410797 LMBRD1 chr6 70411254 70411470 LMBRD1 chr6 70451620 70451836 LMBRD1 chr6 70462044 70462260 LMBRD1 chr6 80816344 80816608 BCKDHB chr6 80837154 80837370 BCKDHB chr6 80838847 80839039 BCKDHB chr6 80877344 80877512 BCKDHB chr6 80878538 80878802 BCKDHB chr6 80880947 80881163 BCKDHB chr6 80910607 80910799 BCKDHB chr6 80912768 80912936 BCKDHB chr6 80982799 80982991 BCKDHB chr6 81053358 81053550 BCKDHB chr6 131894368 131894536 ARG1 chr6 131897743 131897935 ARG1 chr6 131900183 131900447 ARG1 chr6 131902310 131902574 ARG1 chr6 131903652 131903868 ARG1 chr6 131904173 131904389 ARG1 chr6 131904460 131904676 ARG1 chr6 131904831 131905095 ARG1 chr6 150710561 150710705 IYD chr6 150715254 150715470 IYD chr6 158532339 158532555 SERAC1 chr6 158534411 158534627 SERAC1 chr6 158535805 158536021 SERAC1 chr6 158537151 158537343 SERAC1 chr6 158538703 158538871 SERAC1 chr6 158540093 158540309 SERAC1 chr6 158541415 158541655 SERAC1 chr6 158549131 158549347 SERAC1 chr6 158564060 158564276 SERAC1 chr6 158565339 158565555 SERAC1 chr6 158567753 158567969 SERAC1 chr6 158571434 158571674 SERAC1 chr6 158576489 158576681 SERAC1 chr6 158579241 158579433 SERAC1 chr7 40277155 40277371 SUGCT chr7 65425908 65426100 GUSB chr7 65429320 65429488 GUSB chr7 65432701 65432941 GUSB chr7 65435208 65435424 GUSB chr7 65439281 65439425 GUSB chr7 65439479 65439743 GUSB chr7 65439854 65440070 GUSB chr7 65440963 65441107 GUSB chr7 65444356 65444572 GUSB chr7 65444711 65444951 GUSB chr7 65445230 65445446 GUSB chr7 65447007 65447223 GUSB chr7 65546739 65547003 ASL chr7 65547282 65547498 ASL chr7 65547810 65547978 ASL chr7 65548023 65548215 ASL chr7 65551570 65551810 ASL chr7 65552272 65552416 ASL chr7 65552680 65552824 ASL chr7 65553753 65553897 ASL chr7 65554051 65554195 ASL chr7 65554208 65554376 ASL chr7 65554547 65554739 ASL chr7 65556981 65557125 ASL chr7 65557493 65557923 ASL chr7 95750507 95750723 SLC25A13 chr7 95750937 95751105 SLC25A13 chr7 95751129 95751369 SLC25A13 chr7 95761086 95761302 SLC25A13 chr7 95775893 95776037 SLC25A13 chr7 95799301 95799493 SLC25A13 chr7 95800710 95800926 SLC25A13 chr7 95813533 95813773 SLC25A13 chr7 95814217 95814361 SLC25A13 chr7 95818578 95818794 SLC25A13 chr7 95818832 95819024 SLC25A13 chr7 95820417 95820561 SLC25A13 chr7 95822295 95822535 SLC25A13 chr7 95838097 95838313 SLC25A13 chr7 95864005 95864221 SLC25A13 chr7 95906543 95906759 SLC25A13 chr7 95951153 95951369 SLC25A13 chr7 107531649 107531793 DLD chr7 107533634 107533778 DLD chr7 107542150 107542318 DLD chr7 107542710 107542854 DLD chr7 107543820 107544036 DLD chr7 107545297 107545513 DLD chr7 107546701 107546869 DLD chr7 107555903 107556191 DLD chr7 107557256 107557400 DLD chr7 107557698 107557962 DLD chr7 107558364 107558580 DLD chr7 107559407 107559599 DLD chr7 107559602 107559746 DLD chr7 107571829 107572045 LAMB1 chr7 117120085 117120253 CFTR chr7 117144256 117144472 CFTR chr7 117149035 117149251 CFTR chr7 117170939 117171179 CFTR chr7 117174268 117174484 CFTR chr7 117175252 117175516 CFTR chr7 117176577 117176817 CFTR chr7 117178935 117179151 CFTR chr7 117180145 117180409 CFTR chr7 117182007 117182223 CFTR chr7 117188664 117188976 CFTR chr7 117199458 117199770 CFTR chr7 117227729 117227945 CFTR chr7 117229418 117229634 CFTR chr7 117230356 117230548 CFTR chr7 117231978 117232722 CFTR chr7 117234928 117235168 CFTR chr7 117242828 117242972 CFTR chr7 117243579 117243843 CFTR chr7 117246665 117246857 CFTR chr7 117250516 117250780 CFTR chr7 117251606 117252062 CFTR chr7 117252117 117252285 CFTR chr7 117254611 117254827 CFTR chr7 117267563 117267875 CFTR chr7 117279907 117280123 CFTR chr7 117282462 117283254 CFTR chr7 117292845 117293037 CFTR chr7 117304737 117305625 CFTR chr7 117306958 117307198 CFTR chr8 17914959 17915175 AS A FI 1 chr8 17916243 17916459 AS A FI 1 chr8 17916841 17916985 AS A FI 1 chr8 17917025 17917217 AS A FI 1 chr8 17918851 17919019 AS A FI 1 chr8 17919142 17919286 AS A FI 1 chr8 17919727 17919991 AS A FI 1 chr8 17920674 17920818 AS A FI 1 chr8 17921910 17922102 AS A FI 1 chr8 17924620 17924836 AS A FI 1 chr8 17927263 17927431 AS A FI 1 chr8 17928791 17928935 AS A FI 1 chr8 17932994 17933138 AS A FI 1 chr8 17941389 17941605 AS A FI 1 chr8 38001788 38001956 STAR chr8 38002697 38002889 STAR chr8 38003422 38003686 STAR chr8 38005666 38005906 STAR chr8 38006102 38006318 STAR chr8 38008164 38008380 STAR chr8 43002088 43002256 HGSNAT chr8 43014006 43014246 HGSNAT chr8 43016501 43016717 HGSNAT chr8 43024251 43024467 HGSNAT chr8 43025728 43025944 HGSNAT chr8 43027400 43027544 HGSNAT chr8 43028771 43028987 HGSNAT chr8 43033163 43033379 HGSNAT chr8 43037243 43037435 HGSNAT chr8 43046557 43046797 HGSNAT chr8 43047441 43047657 HGSNAT chr8 43048876 43049044 HGSNAT chr8 43052085 43052229 HGSNAT chr8 43052717 43053121 HGSNAT chr8 43054530 43054722 HGSNAT chr8 110099682 110099898 TRHR chr8 110099926 110100190 TRHR chr8 133880269 133880485 TG chr8 133881953 133882169 TG chr8 133883482 133883698 TG chr8 133885199 133885575 TG chr8 133894746 133894962 TG chr8 133898642 133898810 TG chr8 133898851 133899067 TG chr8 133899127 133899343 TG chr8 133900357 133900549 TG chr8 133900643 133900859 TG chr8 133910946 133911162 TG chr8 133913633 133913777 TG chr8 133918977 133919145 TG chr8 133923568 133923784 TG chr8 133935534 133935750 TG chr8 133953630 133953846 TG chr8 133961065 133961281 TG chr8 133975177 133975393 TG chr8 133978835 133979051 TG chr8 133979934 133980263 TG chr8 133981626 133981842 TG chr8 133983942 133984158 TG chr8 133995486 133995702 TG chr8 134024154 134024370 TG chr8 134030077 134030293 TG chr8 134034265 134034481 TG chr8 134042044 134042260 TG chr8 134146826 134147042 TG chr8 143955718 143955934 CYP11B1 chr8 143956312 143956780 CYP11B1 chr8 143957075 143957363 CYP11B1 chr8 143957599 143957839 CYP11B1 chr8 143958043 143958211 CYP11B1 chr8 143958323 143958539 CYP11B1 chr8 143958550 143958694 CYP11B1 chr8 143960390 143960654 CYP11B1 chr8 143960955 143961243 CYP11B1 chr8 145109484 145109652 OPLAH chr8 145109930 145110146 OPLAH chr8 145112924 145113140 OPLAH chr8 145113627 145113819 OPLAH chr9 6532990 6533182 GLDC chr9 6534650 6534842 GLDC chr9 6536004 6536292 GLDC chr9 6540000 6540192 GLDC chr9 6550776 6550968 GLDC chr9 6553320 6553560 GLDC chr9 6554617 6554833 GLDC chr9 6556109 6556325 GLDC chr9 6558499 6558739 GLDC chr9 6565297 6565465 GLDC chr9 6587084 6587324 GLDC chr9 6588309 6588525 GLDC chr9 6588575 6588743 GLDC chr9 6589141 6589285 GLDC chr9 6592131 6592275 GLDC chr9 6592796 6593036 GLDC chr9 6595023 6595167 GLDC chr9 6602067 6602259 GLDC chr9 6604529 6604817 GLDC chr9 6605081 6605249 GLDC chr9 6606487 6606703 GLDC chr9 6610166 6610430 GLDC chr9 6620139 6620379 GLDC chr9 6644505 6644721 GLDC chr9 6645166 6645589 GLDC chr9 34646648 34646840 GALT chr9 34647026 34647314 GALT chr9 34647475 34647715 GALT chr9 34647774 34648014 GALT chr9 34648057 34648225 GALT chr9 34648272 34648512 GALT chr9 34648752 34649088 GALT chr9 34649355 34649667 GALT chr9 34650310 34650502 GALT chr9 93976600 93976768 AUH chr9 93978281 93978497 AUH chr9 93979479 93979695 AUH chr9 93983051 93983243 AUH chr9 94058251 94058419 AUH chr9 94060218 94060362 AUH chr9 94118102 94118318 AUH chr9 94118331 94118547 AUH chr9 94123926 94124142 AUH chr9 100616308 100616608 F0XE1 chr9 104184040 104184232 ALDOB chr9 104187111 104187375 ALDOB chr9 104187755 104187971 ALDOB chr9 104188798 104188966 ALDOB chr9 104189716 104189980 ALDOB chr9 104190694 104190862 ALDOB chr9 104191983 104192127 ALDOB chr9 104192132 104192300 ALDOB chr9 104192949 104193268 ALDOB chr9 104197882 104198098 ALDOB chr9 119460086 119460278 ASTN2 chr9 119460301 119460780 ASTN2 chr9 119460830 119461046 ASTN2 chr9 119461079 119461271 ASTN2 chr9 119461333 119461645 ASTN2 chr9 119461653 119461845 ASTN2 chr9 133327560 133327728 ASS1 chr9 133329650 133329818 ASS1 chr9 133333734 133334022 ASS1 chr9 133339444 133339612 ASS1 chr9 133342058 133342226 ASS1 chr9 133346166 133346358 ASS1 chr9 133346766 133346982 ASS1 chr9 133352196 133352412 ASS1 chr9 133355049 133355289 ASS1 chr9 133355720 133355888 ASS1 chr9 133364668 133364908 ASS1 chr9 133370200 133370464 ASS1 chr9 133374832 133375000 ASS1 chr9 133376254 133376470 ASS1 chr9 139089385 139089601 LHX3 chr9 139090551 139090695 LHX3 chr9 139091407 139091742 LHX3 chr9 139092493 139092637 LHX3 chrlO 6061318 6061534 IL2RA chrlO 6063419 6063766 IL2RA chrlO 6066165 6066381 IL2RA chrlO 6067694 6068050 IL2RA chrlO 6103944 6104160 IL2RA chrlO 14950421 14951434 DCLRE1C chrlO 14968792 14968936 DCLRE1C chrlO 14969906 14970122 DCLRE1C chrlO 14974794 14974938 DCLRE1C chrlO 14976373 14976517 DCLRE1C chrlO 14976666 14976810 DCLRE1C chrlO 14977428 14977572 DCLRE1C chrlO 14978434 14978650 DCLRE1C chrlO 14987079 14987223 DCLRE1C chrlO 14995850 14996066 DCLRE1C chrlO 69556815 69557007 DNAJC12 chrlO 69560017 69560425 DNAJC12 chrlO 69560703 69561279 DNAJC12 chrlO 69561897 69562065 DNAJC12 chrlO 69562091 69562259 DNAJC12 chrlO 69562373 69562589 DNAJC12 chrlO 69563017 69564049 DNAJC12 chrlO 69564346 69564514 DNAJC12 chrlO 69564789 69564957 DNAJC12 chrlO 69565091 69565571 DNAJC12 chrlO 69565599 69565815 DNAJC12 chrlO 69565843 69566299 DNAJC12 chrlO 69571310 69571478 DNAJC12 chrlO 69582989 69583205 DNAJC12 chrlO 69597604 69597820 DNAJC12 chrlO 72643652 72643844 PCBD1 chrlO 72645556 72645700 PCBD1 chrlO 75960437 75960653 ADK chrlO 76285054 76285270 ADK chrlO 76348946 76349162 ADK chrlO 76360080 76360296 ADK chrlO 76429908 76430124 ADK chrlO 76468037 76468253 ADK chrlO 82033485 82033701 MAT1A chrlO 82034235 82034451 MAT1A chrlO 82034712 82035049 MAT1A chrlO 82036029 82036245 MAT1A chrlO 82039898 82040042 MAT1A chrlO 82043564 82043780 MAT1A chrlO 82045165 82045381 MAT1A chrlO 90974573 90974861 LI PA chrlO 90975649 90975793 LI PA chrlO 90982211 90982355 LI PA chrlO 90983382 90983692 LI PA chrlO 90984815 90984983 LI PA chrlO 90986614 90986806 LI PA chrlO 90987900 90988188 LI PA chrlO 91005417 91005585 LI PA chrlO 91007186 91007513 LI PA chrlO 91011384 91011600 LI PA chrlO 104590487 104590751 CYP17A1 chrlO 104591207 104591399 CYP17A1 chrlO 104592236 104592476 CYP17A1 chrlO 104592731 104592899 CYP17A1 chrlO 104593779 104593923 CYP17A1 chrlO 104594499 104594881 CYP17A1 chrlO 104594956 104595196 CYP17A1 chrlO 104596813 104597125 CYP17A1 chrlO 124797253 124797469 ACADSB chrlO 124799945 124800161 ACADSB chrlO 124800779 124800923 ACADSB chrlO 124802506 124802698 ACADSB chrlO 124812561 124812729 ACADSB chrlO 126086470 126086710 OAT chrlO 126089395 126089587 OAT chrlO 126090244 126090460 OAT chrlO 126091440 126091680 OAT chrlO 126092329 126092521 OAT chrlO 126093977 126094169 OAT chrlO 126097094 126097262 OAT chrlO 126097342 126097582 OAT chrlO 126100481 126100793 OAT chrll 5246616 5247240 HBB chrll 5247799 5248783 HBB chrll 6411733 6411949 SMPD1 chrll 6411952 6412240 SMPD1 chrll 6412638 6413382 SMPD1 chrll 6414394 6414658 SMPD1 chrll 6414795 6414963 SMPD1 chrll 6415060 6415300 SMPD1 chrll 6415420 6415780 SMPD1 chrll 6635734 6635974 TPP1 chrll 6636047 6636263 TPP1 chrll 6636351 6636854 TPP1 chrll 6637175 6637367 TPP1 chrll 6637542 6637734 TPP1 chrll 6637857 6638145 TPP1 chrll 6638151 6638439 TPP1 chrll 6638479 6638719 TPP1 chrll 6638798 6639062 TPP1 chrll 6639959 6640103 TPP1 chrll 6640314 6640722 TPP1 chrll 36595103 36595415 RAG1 chrll 36595543 36595687 RAG1 chrll 36595742 36595982 RAG1 chrll 36596020 36596548 RAG1 chrll 36596613 36596829 RAG1 chrll 36596851 36597211 RAG1 chrll 36597284 36597476 RAG1 chrll 36597528 36597840 RAG1 chrll 36614260 36614452 RAG2 chrll 36614707 36614923 RAG2 chrll 36615089 36615826 RAG2 chrll 68525114 68525258 CPT1A chrll 68527008 68527224 CPT1A chrll 68527656 68527824 CPT1A chrll 68528945 68529209 CPT1A chrll 68530070 68530286 CPT1A chrll 68540677 68540958 CPT1A chrll 68542734 68542950 CPT1A chrll 68548048 68548264 CPT1A chrll 68549185 68549425 CPT1A chrll 68552231 68552495 CPT1A chrll 68560726 68560894 CPT1A chrll 68562273 68562489 CPT1A chrll 68564267 68564459 CPT1A chrll 68566577 68566793 CPT1A chrll 68571420 68571612 CPT1A chrll 68574991 68575207 CPT1A chrll 68579844 68580060 CPT1A chrll 68582762 68582906 CPT1A chrll 71146415 71146895 DHCR7 chrll 71148803 71149043 DHCR7 chrll 71149871 71150183 DHCR7 chrll 71152260 71152500 DHCR7 chrll 71153245 71153461 DHCR7 chrll 71154989 71155133 DHCR7 chrll 71155151 71155319 DHCR7 chrll 71155839 71156055 DHCR7 chrll 107992278 107992470 ACAT1 chrll 108002611 108002827 ACAT1 chrll 108004481 108004697 ACAT1 chrll 108004931 108005099 ACAT1 chrll 108005833 108006025 ACAT1 chrll 108009580 108009820 ACAT1 chrll 108010781 108010997 ACAT1 chrll 108012227 108012491 ACAT1 chrll 108013084 108013300 ACAT1 chrll 108014650 108014818 ACAT1 chrll 108016875 108017139 ACAT1 chrll 108017944 108018136 ACAT1 chrll 112097146 112097290 PTS chrll 112098938 112099082 PTS chrll 112099257 112099449 PTS chrll 112099909 112100125 PTS chrll 112100870 112101014 PTS chrll 112101312 112101456 PTS chrll 112103825 112104017 PTS chrll 112104104 112104296 PTS chrll 118182778 118182994 CD3E chrll 118183303 118183543 CD3E chrll 118184503 118184647 CD3E chrll 118209822 118210038 CD3D chrll 118210509 118210725 CD3D chrll 118211029 118211293 CD3D chrll 134123437 134123605 ACAD8 chrll 134126357 134126573 ACAD8 chrll 134126896 134127227 ACAD8 chrll 134128374 134128566 ACAD8 chrll 134128871 134129087 ACAD8 chrll 134130854 134131070 ACAD8 chrll 134131086 134131302 ACAD8 chrll 134132400 134132568 ACAD8 chrl2 103232903 103233047 PAH chrl2 103234117 103234357 PAH chrl2 103237356 103237620 PAH chrl2 103238044 103238260 PAH chrl2 103240619 103240787 PAH chrl2 103245403 103245595 PAH chrl2 103246532 103246796 PAH chrl2 103248172 103248597 PAH chrl2 103248613 103249117 PAH chrl2 103260314 103260506 PAH chrl2 103271186 103271378 PAH chrl2 103288454 103288766 PAH chrl2 103306512 103306680 PAH chrl2 103310792 103310960 PAH chrl2 103351930 103352146 ASCL1 chrl2 109994834 109994978 MMAB chrl2 109996847 109997015 MMAB chrl2 109998811 109998979 MMAB chrl2 109999173 109999341 MMAB chrl2 109999572 109999716 MMAB chrl2 110002888 110003032 MMAB chrl2 110006527 110006719 MMAB chrl2 110011124 110011340 MMAB chrl2 110012564 110012756 MVK chrl2 110013744 110014008 MVK chrl2 110017597 110017789 MVK chrl2 110019144 110019408 MVK chrl2 110023791 110023983 MVK chrl2 110024458 110024674 MVK chrl2 110028527 110028719 MVK chrl2 110029015 110029159 MVK chrl2 110032799 110033039 MVK chrl2 110034224 110034440 MVK chrl2 121163634 121163778 ACADS chrl2 121164832 121165048 ACADS chrl2 121174732 121174996 ACADS chrl2 121175107 121175275 ACADS chrl2 121175580 121175820 ACADS chrl2 121176027 121176195 ACADS chrl2 121176293 121176509 ACADS chrl2 121176607 121176775 ACADS chrl2 121176887 121177055 ACADS chrl2 121177067 121177283 ACADS chrl2 122277557 122277968 HPD chrl2 122284717 122284933 HPD chrl2 122285009 122285225 HPD chrl2 122287524 122287740 HPD chrl2 122295182 122295398 HPD chrl2 122295625 122295769 HPD chrl3 20763049 20763745 GJB2 chrl3 20766739 20766955 GJB2 chrl3 41367275 41367491 SLC25A15 chrl3 41373140 41373404 SLC25A15 chrl3 41379227 41379443 SLC25A15 chrl3 41381435 41381603 SLC25A15 chrl3 41382495 41382711 SLC25A15 chrl3 41383656 41383800 SLC25A15 chrl3 48523017 48523233 SUCLA2 chrl3 48528490 48528730 SUCLA2 chrl3 48542674 48542890 SUCLA2 chrl3 48562619 48562787 SUCLA2 chrl3 48562981 48563173 SUCLA2 chrl3 48877878 48878094 RBI chrl3 48878109 48878277 RBI chrl3 48881349 48881589 RBI chrl3 48916689 48916905 RBI chrl3 48919156 48919396 RBI chrl3 48921912 48922056 RBI chrl3 48923047 48923215 RBI chrl3 48934138 48934306 RBI chrl3 48936890 48937154 RBI chrl3 48938973 48939165 RBI chrl3 48941578 48941794 RBI chrl3 48942603 48942747 RBI chrl3 48947489 48947681 RBI chrl3 48951004 48951220 RBI chrl3 48953667 48953835 RBI chrl3 48954274 48954466 RBI chrl3 48955326 48955638 RBI chrl3 48985633 48985849 RBI chrl3 48985962 48986202 RBI chrl3 48986264 48986480 RBI chrl3 49027064 49027304 RBI chrl3 49030283 49030547 RBI chrl3 49033765 49034029 RBI chrl3 49037811 49038027 RBI chrl3 49039130 49039514 RBI chrl3 49045990 49046206 RBI chrl3 49047441 49047585 RBI chrl3 49050775 49051088 RBI chrl3 49051411 49051627 RBI chrl3 52508891 52509179 ATP7B chrl3 52509673 52509889 ATP7B chrl3 52511410 52511818 ATP7B chrl3 52513137 52513353 ATP7B chrl3 52515163 52515403 ATP7B chrl3 52516480 52516744 ATP7B chrl3 52518238 52518982 ATP7B chrl3 52520378 52520666 ATP7B chrl3 52523746 52523986 ATP7B chrl3 52524089 52524564 ATP7B chrl3 52531601 52531793 ATP7B chrl3 52532436 52532700 ATP7B chrl3 52534223 52534535 ATP7B chrl3 52535896 52536112 ATP7B chrl3 52538980 52539244 ATP7B chrl3 52542511 52542799 ATP7B chrl3 52544575 52544887 ATP7B chrl3 52548067 52548835 ATP7B chrl3 52548951 52549311 ATP7B chrl3 52585361 52585529 ATP7B chrl3 52585796 52586012 ATP7B chrl3 100741320 100741536 PCCA chrl3 100755105 100755273 PCCA chrl3 100764035 100764357 PCCA chrl3 100807183 100807399 PCCA chrl3 100809482 100809650 PCCA chrl3 100861555 100861771 PCCA chrl3 100888002 100888218 PCCA chrl3 100909811 100909955 PCCA chrl3 100914963 100915107 PCCA chrl3 100920907 100921075 PCCA chrl3 100925393 100925657 PCCA chrl3 100953655 100953919 PCCA chrl3 100955137 100955353 PCCA chrl3 100959401 100959569 PCCA chrl3 100962043 100962259 PCCA chrl3 100982819 100983035 PCCA chrl3 100992383 100992599 PCCA chrl3 101020680 101020896 PCCA chrl3 101077827 101078043 PCCA chrl3 101101447 101101567 PCCA chrl3 101167651 101167891 PCCA chrl3 101179871 101180063 PCCA chrl3 101182286 101182454 PCCA chrl3 108860920 108861590 LIG4 chrl3 108861605 108862581 LIG4 chrl3 108862656 108862872 LIG4 chrl3 108862896 108863112 LIG4 chrl3 108863305 108863521 LIG4 chrl4 20940417 20940740 PNP chrl4 20942617 20943140 PNP chrl4 20943223 20943391 PNP chrl4 20944540 20944732 PNP chrl4 23242783 23242951 SLC7A7 chrl4 23243100 23243364 SLC7A7 chrl4 23243525 23243741 SLC7A7 chrl4 23244591 23244807 SLC7A7 chrl4 23244987 23245203 SLC7A7 chrl4 23245352 23245568 SLC7A7 chrl4 23247967 23248111 SLC7A7 chrl4 23249078 23249270 SLC7A7 chrl4 23282106 23282610 SLC7A7 chrl4 36986375 36986732 NKX2-1 chrl4 36986748 36986892 NKX2-1 chrl4 36986898 36987234 NKX2-1 chrl4 36988138 36988450 NKX2-1 chrl4 36989226 36989442 NKX2-1 chrl4 55310394 55310610 GCH1 chrl4 55310677 55310917 GCH1 chrl4 55312427 55312619 GCH1 chrl4 55313761 55313905 GCH1 chrl4 55326344 55326488 GCH1 chrl4 55332015 55332207 GCH1 chrl4 55369052 55369388 GCH1 chrl4 74753098 74753314 ABCD4 chrl4 74753351 74753495 ABCD4 chrl4 74754453 74754669 ABCD4 chrl4 74754843 74755059 ABCD4 chrl4 74759218 74759434 ABCD4 chrl4 74762927 74763143 ABCD4 chrl4 74946884 74947100 NPC2 chrl4 74947351 74947543 NPC2 chrl4 74951063 74951351 NPC2 chrl4 74953014 74953158 NPC2 chrl4 74959840 74960032 NPC2 chrl4 81421987 81422203 TSHR chrl4 81534516 81534732 TSHR chrl4 81554198 81554414 TSHR chrl4 81557330 81557546 TSHR chrl4 81558839 81559007 TSHR chrl4 81562877 81563093 TSHR chrl4 81606015 81606231 TSHR chrl4 81609279 81609423 TSHR chrl4 81609571 81609979 TSHR chrl4 81610189 81610429 TSHR chrl4 88401038 88401278 GALC chrl4 88406189 88406381 GALC chrl4 88407699 88407963 GALC chrl4 88411842 88412082 GALC chrl4 88414011 88414227 GALC chrl4 88416139 88416307 GALC chrl4 88416960 88417176 GALC chrl4 88429665 88429881 GALC chrl4 88431800 88431992 GALC chrl4 88434623 88434815 GALC chrl4 88442646 88442886 GALC chrl4 88448494 88448638 GALC chrl4 88450691 88450931 GALC chrl4 88452780 88452996 GALC chrl4 88454379 88454595 GALC chrl4 88454749 88454917 GALC chrl4 88459251 88459563 GALC chrl4 88459667 88459883 GALC chrl4 94844811 94845915 SERPINA1 chrl4 94847249 94847489 SERPINA1 chrl4 94848874 94849090 SERPINA1 chrl4 94849151 94849415 SERPINA1 chrl4 94849456 94849672 SERPINA1 chrl5 40697965 40698229 IVD chrl5 40699786 40699978 IVD chrl5 40700038 40700254 IVD chrl5 40702818 40703562 IVD chrl5 40703695 40703911 IVD chrl5 40705154 40705346 IVD chrl5 40707039 40707231 IVD chrl5 40707547 40707739 IVD chrl5 40708216 40708384 IVD chrl5 40708440 40708608 IVD chrl5 40710309 40710501 IVD chrl5 45386331 45386547 DU0X2 chrl5 45389326 45389542 DU0X2 chrl5 45389736 45389952 DU0X2 chrl5 45390149 45390365 DU0X2 chrl5 45391488 45391790 DU0X2 chrl5 45391884 45392076 DU0X2 chrl5 45392934 45393150 DU0X2 chrl5 45393315 45393531 DU0X2 chrl5 45394069 45394285 DU0X2 chrl5 45396051 45396267 DU0X2 chrl5 45396314 45396530 DU0X2 chrl5 45398307 45398633 DU0X2 chrl5 45398680 45398896 DU0X2 chrl5 45399540 45399756 DU0X2 chrl5 45400241 45400457 DU0X2 chrl5 45400977 45401193 DU0X2 chrl5 45402041 45402185 DU0X2 chrl5 45402580 45402796 DU0X2 chrl5 45403587 45403803 DU0X2 chrl5 45403906 45404194 DU0X2 chrl5 45405129 45405345 DU0X2 chrl5 45406703 45406919 DU0X2 chrl5 45408678 45408894 DU0XA2 chrl5 45409364 45409580 DU0XA2 chrl5 45654262 45654478 GATM chrl5 45656038 45656254 GATM chrl5 45658496 45658712 GATM chrl5 45660228 45660546 GATM chrl5 45661471 45661615 GATM chrl5 45668763 45668979 GATM chrl5 76508831 76509047 ETFA chrl5 76566664 76566880 ETFA chrl5 76576056 76576272 ETFA chrl5 76577926 76578070 ETFA chrl5 76578719 76578863 ETFA chrl5 76584669 76584885 ETFA chrl5 76587958 76588174 ETFA chrl5 76603618 76603834 ETFA chrl5 80445336 80445528 FAH chrl5 80450294 80450621 FAH chrl5 80452038 80452278 FAH chrl5 80452693 80452861 FAH chrl5 80454567 80454735 FAH chrl5 80460382 80460670 FAH chrl5 80464433 80464649 FAH chrl5 80465301 80465541 FAH chrl5 80467312 80467456 FAH chrl5 80469818 80470034 FAH chrl5 80472410 80472626 FAH chrl5 80473322 80473562 FAH chrl5 80478414 80478606 FAH chrl6 71602055 71602223 TAT chrl6 71603700 71603892 TAT chrl6 71604062 71604278 TAT chrl6 71604517 71604733 TAT chrl6 71605454 71605670 TAT chrl6 71606019 71606337 TAT chrl6 71606440 71606656 TAT chrl6 71609716 71610201 TAT chrl6 71610217 71610433 TAT chrl6 83932632 83932800 MLYCD chrl6 83932836 83933052 MLYCD chrl6 83933114 83933330 MLYCD chrl6 83940567 83940759 MLYCD chrl6 83941664 83941880 MLYCD chrl6 83945862 83946078 MLYCD chrl6 83948492 83948732 MLYCD chrl6 83948799 83949015 MLYCD chrl6 88880798 88880990 GALNS chrl6 88884365 88884533 GALNS chrl6 88888888 88889104 GALNS chrl6 88891137 88891329 GALNS chrl6 88893050 88893290 GALNS chrl6 88898360 88898624 GALNS chrl6 88901565 88901805 GALNS chrl6 88902102 88902294 GALNS chrl6 88902527 88902743 GALNS chrl6 88903988 88904228 GALNS chrl6 88904927 88905143 GALNS chrl6 88907349 88907541 GALNS chrl6 88908239 88908455 GALNS chrl6 88909076 88909292 GALNS chrl6 88923134 88923326 GALNS chrl7 3543428 3543620 CTNS chrl7 3550681 3550873 CTNS chrl7 3552143 3552287 CTNS chrl7 3558261 3558453 CTNS chrl7 3558518 3558686 CTNS chrl7 3559786 3560098 CTNS chrl7 3561246 3561510 CTNS chrl7 3563101 3563317 CTNS chrl7 3563470 3563710 CTNS chrl7 7120385 7120601 DLG4 chrl7 7121015 7121183 DLG4 chrl7 7123256 7123568 ACADVL chrl7 7123734 7124046 ACADVL chrl7 7124098 7124362 ACADVL chrl7 7124799 7125039 ACADVL chrl7 7125221 7125437 ACADVL chrl7 7125445 7125661 ACADVL chrl7 7125931 7126243 ACADVL chrl7 7126418 7126610 ACADVL chrl7 7126941 7127397 ACADVL chrl7 7127447 7127615 ACADVL chrl7 7127632 7127896 ACADVL chrl7 7127949 7128189 ACADVL chrl7 7128237 7128405 ACADVL chrl7 38233665 38233881 THRA chrl7 38242916 38243132 THRA chrl7 38244451 38244667 THRA chrl7 38245527 38245743 THRA chrl7 40688209 40688353 NAGLU chrl7 40688536 40688752 NAGLU chrl7 40689358 40689622 NAGLU chrl7 40690306 40690577 NAGLU chrl7 40690644 40690812 NAGLU chrl7 40692947 40693259 NAGLU chrl7 40695085 40695733 NAGLU chrl7 40695828 40696260 NAGLU chrl7 42081981 42082149 PYY chrl7 42082347 42082563 NAGS chrl7 42083090 42083258 NAGS chrl7 42083417 42083633 NAGS chrl7 42083842 42084058 NAGS chrl7 42084756 42085248 NAGS chrl7 42085808 42086024 NAGS chrl7 73748194 73748612 ITGB4 chrl7 73750647 73750863 ITGB4 chrl7 73751780 73751996 ITGB4 chrl7 73753096 73753240 ITGB4 chrl7 73753254 73753625 ITGB4,GALK1 chrl7 73754064 73754280 GALK1 chrl7 73754297 73754441 GALK1 chrl7 73754479 73754671 GALK1 chrl7 73758758 73759022 GALK1 chrl7 73759358 73759574 GALK1 chrl7 73759981 73760149 GALK1 chrl7 73761031 73761295 GALK1 chrl7 78078336 78078936 GAA chrl7 78079514 78079754 GAA chrl7 78081281 78081793 GAA chrl7 78082108 78082636 GAA chrl7 78083694 78083910 GAA chrl7 78084511 78084823 GAA chrl7 78085724 78085964 GAA chrl7 78086327 78086543 GAA chrl7 78086622 78086886 GAA chrl7 78086975 78087215 GAA chrl7 78090729 78090969 GAA chrl7 78091390 78092614 GAA chrl7 78093032 78093176 GAA chrl7 78184271 78184799 SGSH chrl7 78185850 78186114 SGSH chrl7 78187566 78187734 SGSH chrl7 78187952 78188144 SGSH chrl7 78188378 78188618 SGSH chrl7 78188819 78188987 SGSH chrl7 78190787 78191051 SGSH chrl7 78194002 78194170 SGSH chrl8 21112103 21112319 NPC1 chrl8 21113268 21113532 NPC1 chrl8 21114350 21114566 NPC1 chrl8 21115429 21115669 NPC1 chrl8 21116587 21116899 NPC1 chrl8 21118471 21118687 NPC1 chrl8 21119265 21119481 NPC1 chrl8 21119718 21120006 NPC1 chrl8 21120357 21120525 NPC1 chrl8 21120976 21121192 NPC1 chrl8 21121221 21121413 NPC1 chrl8 21123337 21123553 NPC1 chrl8 21124257 21124545 NPC1 chrl8 21124872 21125160 NPC1 chrl8 21127921 21128065 NPC1 chrl8 21128272 21128488 NPC1 chrl8 21128568 21128688 NPC1 chrl8 21130260 21130380 NPC1 chrl8 21130787 21131963 NPC1 chrl8 21131975 21132143 NPC1 chrl8 21132259 21133387 NPC1 chrl8 21134716 21134956 NPC1 chrl8 21136220 21136580 NPC1 chrl8 21140183 21140423 NPC1 chrl8 21141271 21141511 NPC1 chrl8 21148739 21149003 NPC1 chrl8 21152029 21152245 NPC1 chrl8 21153358 21153598 NPC1 chrl8 21166198 21166414 NPC1 chrl9 1397293 1397509 GAMT chrl9 1398900 1399068 GAMT chrl9 1399086 1399254 GAMT chrl9 1399471 1399639 GAMT chrl9 1399734 1399998 GAMT chrl9 1401258 1401522 GAMT chrl9 12757364 12757580 MAN2B1 chrl9 12757993 12758161 MAN2B1 chrl9 12758203 12758467 MAN2B1 chrl9 12758880 12759096 MAN2B1 chrl9 12759110 12759326 MAN2B1 chrl9 12759894 12760086 MAN2B1 chrl9 12760103 12760295 MAN2B1 chrl9 12760735 12761047 MAN2B1 chrl9 12762965 12763277 MAN2B1 chrl9 12766457 12766745 MAN2B1 chrl9 12767390 12767558 MAN2B1 chrl9 12767708 12767876 MAN2B1 chrl9 12768227 12768419 MAN2B1 chrl9 12768819 12768963 MAN2B1 chrl9 12769054 12769294 MAN2B1 chrl9 12771969 12772185 MAN2B1 chrl9 12774193 12774337 MAN2B1 chrl9 12774456 12774648 MAN2B1 chrl9 12775584 12775898 MAN2B1 chrl9 12776125 12776433 MAN2B1 chrl9 12776445 12776661 MAN2B1 chrl9 12777303 12777567 MAN2B1 chrl9 13002071 13002263 GCDH chrl9 13002608 13002848 GCDH chrl9 13002876 13003044 GCDH chrl9 13004243 13004531 GCDH chrl9 13006759 13006927 GCDH chrl9 13007003 13007243 GCDH chrl9 13007669 13007885 GCDH chrl9 13008104 13008296 GCDH chrl9 13008469 13008733 GCDH chrl9 13010222 13010414 GCDH chrl9 17937592 17937808 JAK3 chrl9 17940914 17941130 JAK3 chrl9 17941311 17941527 JAK3 chrl9 17941997 17942213 JAK3 chrl9 17942487 17942703 JAK3 chrl9 17945299 17945515 JAK3 chrl9 17945609 17945825 JAK3 chrl9 17945862 17946078 JAK3 chrl9 17946675 17946867 JAK3 chrl9 17947899 17948067 JAK3 chrl9 17948688 17948856 JAK3 chrl9 17949024 17949192 JAK3 chrl9 17950237 17950453 JAK3 chrl9 17950993 17951209 JAK3 chrl9 17952463 17952631 JAK3 chrl9 17953200 17953416 JAK3 chrl9 17953818 17954034 JAK3 chrl9 17954152 17954368 JAK3 chrl9 17954535 17954727 JAK3 chrl9 17954999 17955167 JAK3 chrl9 17983297 17983513 SLC5A5 chrl9 17988533 17988749 SLC5A5 chrl9 17992662 17993077 SLC5A5 chrl9 17999152 17999296 SLC5A5 chrl9 41903682 41903898 BCKDHA chrl9 41916487 41916962 BCKDHA chrl9 41919924 41920116 BCKDHA chrl9 41925005 41925245 BCKDHA chrl9 41928016 41928328 BCKDHA chrl9 41928482 41928722 BCKDHA chrl9 41928860 41929124 BCKDHA chrl9 41930283 41930547 BCKDHA chrl9 44010917 44011133 ETHE1 chrl9 44012097 44012313 ETHE1 chrl9 44012875 44013019 ETHE1 chrl9 44015530 44015746 ETHE1 chrl9 44030291 44030555 ETHE1 chrl9 44030615 44030831 ETHE1 chrl9 44031194 44031386 ETHE1 chrl9 46032315 46032723 0PA3 chrl9 46056665 46057182 0PA3 chrl9 46087830 46088022 0PA3 chrl9 51848528 51848744 ETFB chrl9 51850152 51850368 ETFB chrl9 51853535 51853751 ETFB chrl9 51856434 51856578 ETFB chrl9 51857363 51857649 ETFB chr20 32878427 32878643 AHCY chr20 32880131 32880323 AHCY chr20 32881781 32881997 AHCY chr20 33516541 33516757 GSS chr20 33519113 33519329 GSS chr20 33519816 33520032 GSS chr20 33523301 33523517 GSS chr20 33524530 33524887 GSS chr20 33530183 33530517 GSS chr20 33539545 33539761 GSS chr20 33543417 33543633 GSS chr20 43248395 43248611 ADA chr20 43248887 43249127 ADA chr20 43249605 43249821 ADA chr20 43251177 43251345 ADA chr20 43251462 43251702 ADA chr20 43252790 43253030 ADA chr20 43254157 43254373 ADA chr20 43255034 43255298 ADA chr20 43257579 43257795 ADA chr20 43264799 43264967 ADA chr20 43280160 43280304 ADA chr21 38126505 38126769 HLCS chr21 38128796 38129084 HLCS chr21 38132021 38132189 HLCS chr21 38137260 38137524 HLCS chr21 38139412 38139628 HLCS chr21 38269094 38269489 HLCS chr21 38302493 38302733 HLCS chr21 38308660 38308876 HLCS chr21 38308906 38309218 HLCS chr21 38309221 38309687 HLCS chr21 38311084 38311300 HLCS chr21 44473913 44474153 CBS chr21 44476859 44477051 CBS chr21 44478256 44478472 CBS chr21 44478891 44479131 CBS chr21 44479274 44479466 CBS chr21 44480509 44480701 CBS chr21 44482365 44482557 CBS chr21 44483007 44483295 CBS chr21 44483960 44484152 CBS chr21 44485258 44485450 CBS chr21 44485489 44485969 CBS chr21 44486296 44486536 CBS chr21 44488564 44488756 CBS chr21 44492042 44492210 CBS chr21 44492213 44492357 CBS chr21 44495766 44495982 CBS chr21 47556812 47557028 FTCD chr21 47569935 47570151 FTCD chr21 47571800 47571944 FTCD chr22 18900897 18901113 PRODH chr22 18905845 18906013 PRODH chr22 18906871 18907087 PRODH chr22 18910309 18910525 PRODH chr22 31006772 31007146 TCN2 chr22 31008921 31009137 TCN2 chr22 31010227 31010578 TCN2 chr22 31011004 31011220 TCN2 chr22 31011273 31011465 TCN2 chr22 31011493 31011817 TCN2 chr22 31013367 31013583 TCN2 chr22 31018868 31019084 TCN2 chr22 51063602 51063890 ARSA chr22 51063951 51064167 ARSA chr22 51064355 51064715 ARSA chr22 51065010 51065490 ARSA chr22 51065582 51065846 ARSA chr22 51065981 51066245 ARSA chrX 22051078 22051294 PHEX chrX 22056524 22056716 PHEX chrX 22065145 22065385 PHEX chrX 22094462 22094654 PHEX chrX 22095585 22095873 PHEX chrX 22108487 22108679 PHEX chrX 22112018 22112258 PHEX chrX 22113377 22113593 PHEX chrX 22115018 22115210 PHEX chrX 22117064 22117328 PHEX chrX 22129524 22129740 PHEX chrX 22132520 22132760 PHEX chrX 22151583 22151799 PHEX chrX 22186375 22186567 PHEX chrX 22196336 22196552 PHEX chrX 22208508 22208676 PHEX chrX 22230965 22231133 PHEX chrX 22237092 22237284 PHEX chrX 22239675 22239915 PHEX chrX 22244509 22244677 PHEX chrX 22245566 22245782 PHEX chrX 22263396 22263588 PHEX chrX 22265911 22266127 PHEX chrX 22266193 22266409 PHEX chrX 30322674 30323058 NR0B1 chrX 30326286 30327390 NR0B1 chrX 31144656 31144872 DMD chrX 31165381 31165645 DMD chrX 31187511 31187775 DMD chrX 31190440 31190584 DMD chrX 31191621 31191765 DMD chrX 31196025 31196265 DMD chrX 31196735 31196975 DMD chrX 31198435 31198651 DMD chrX 31200572 31200788 DMD chrX 31200809 31201073 DMD chrX 31222024 31222288 DMD chrX 31224644 31224836 DMD chrX 31227555 31227867 DMD chrX 31241104 31241320 DMD chrX 31279036 31279180 DMD chrX 31279310 31279526 DMD chrX 31279672 31279888 DMD chrX 31341661 31341829 DMD chrX 31366616 31366808 DMD chrX 31462582 31462798 DMD chrX 31496214 31496502 DMD chrX 31497059 31497275 DMD chrX 31514852 31515116 DMD chrX 31525340 31525628 DMD chrX 31526219 31526435 DMD chrX 31627636 31627852 DMD chrX 31645737 31646025 DMD chrX 31676053 31676293 DMD chrX 31697442 31697754 DMD chrX 31747688 31747904 DMD chrX 31792106 31792370 DMD chrX 31838042 31838210 DMD chrX 31854798 31854990 DMD chrX 31893247 31893559 DMD chrX 31947654 31947894 DMD chrX 31950139 31950403 DMD chrX 31983038 31983254 DMD chrX 31986394 31986658 DMD chrX 32234993 32235233 DMD chrX 32305588 32305876 DMD chrX 32328191 32328431 DMD chrX 32360153 32360441 DMD chrX 32361191 32361455 DMD chrX 32364005 32364245 DMD chrX 32366472 32366688 DMD chrX 32380858 32381122 DMD chrX 32382643 32382883 DMD chrX 32383084 32383372 DMD chrX 32398568 32398856 DMD chrX 32404390 32404630 DMD chrX 32407555 32407843 DMD chrX 32408153 32408345 DMD chrX 32429819 32430083 DMD chrX 32456302 32456566 DMD chrX 32459239 32459479 DMD chrX 32466521 32466809 DMD chrX 32472757 32473045 DMD chrX 32479412 32479628 DMD chrX 32481501 32481765 DMD chrX 32482650 32482842 DMD chrX 32486568 32486880 DMD chrX 32490222 32490486 DMD chrX 32502981 32503269 DMD chrX 32509383 32509647 DMD chrX 32519818 32520010 DMD chrX 32536061 32536301 DMD chrX 32563221 32563509 DMD chrX 32582844 32583204 DMD chrX 32583211 32583427 DMD chrX 32583453 32583957 DMD chrX 32591593 32592021 DMD chrX 32613814 32614054 DMD chrX 32632367 32632631 DMD chrX 32662186 32662540 DMD chrX 32663031 32663319 DMD chrX 32715972 32716188 DMD chrX 32717176 32717464 DMD chrX 32756800 32757016 DMD chrX 32827548 32827788 DMD chrX 32834544 32834808 DMD chrX 32841363 32841555 DMD chrX 32841859 32842075 DMD chrX 32862841 32863033 DMD chrX 32867783 32867999 DMD chrX 33038183 33038375 DMD chrX 33192344 33192560 DMD chrX 33229336 33229480 DMD chrX 38211901 38212093 OTC chrX 38226492 38226732 OTC chrX 38229006 38229198 OTC chrX 38240544 38240736 OTC chrX 38260473 38260737 OTC chrX 38260838 38261054 OTC chrX 38262812 38263052 OTC chrX 38267993 38268281 OTC chrX 38271064 38271304 OTC chrX 38280218 38280386 OTC chrX 53458341 53458557 HSD17B10 chrX 53458691 53458883 HSD17B10 chrX 53458925 53459117 HSD17B10 chrX 53459187 53459403 HSD17B10 chrX 70327654 70327798 IL2RG chrX 70328078 70328246 IL2RG chrX 70328390 70328606 IL2RG chrX 70329053 70329293 IL2RG chrX 70329954 70330194 IL2RG chrX 70330297 70330609 IL2RG chrX 70330690 70330906 IL2RG chrX 70331231 70331447 IL2RG chrX 77227060 77227276 ATP7A chrX 77243625 77243841 ATP7A chrX 77243976 77244325 ATP7A chrX 77244886 77245395 ATP7A chrX 77253940 77254228 ATP7A chrX 77258509 77258797 ATP7A chrX 77264614 77264782 ATP7A chrX 77266617 77266809 ATP7A chrX 77266891 77267207 ATP7A chrX 77268372 77268612 ATP7A chrX 77270164 77270308 ATP7A chrX 77271193 77271433 ATP7A chrX 77275686 77275950 ATP7A chrX 77276442 77276634 ATP7A chrX 77284711 77284999 ATP7A chrX 77286841 77286985 ATP7A chrX 77286987 77287131 ATP7A chrX 77289097 77289361 ATP7A chrX 77294249 77294465 ATP7A chrX 77296142 77296286 ATP7A chrX 77298029 77298341 ATP7A chrX 77298766 77298982 ATP7A chrX 77300916 77301132 ATP7A chrX 77301806 77302022 ATP7A chrX 100652793 100653105 GLA chrX 100653303 100653615 GLA chrX 100653721 100653985 GLA chrX 100655605 100655797 GLA chrX 100656566 100656854 GLA chrX 100658742 100659030 GLA chrX 100662638 100662950 GLA chrX 148564329 148564761 IDS chrX 148568431 148568695 IDS chrX 148568736 148568952 IDS chrX 148571786 148572026 IDS chrX 148577820 148578060 IDS chrX 148579582 148579894 IDS chrX 148582453 148582621 IDS chrX 148584789 148585077 IDS chrX 148585634 148585850 IDS chrX 148586556 148586892 IDS chrX 152954028 152954317 SLC6A8 chrX 152955779 152955971 SLC6A8 chrX 152956706 152956874 SLC6A8 chrX 152956877 152957069 SLC6A8 chrX 152957425 152957593 SLC6A8 chrX 152958522 152958865 SLC6A8 chrX 152958923 152959091 SLC6A8 chrX 152959308 152959524 SLC6A8 chrX 152959527 152959695 SLC6A8 chrX 152959789 152960101 SLC6A8 chrX 152960123 152960363 SLC6A8 chrX 152960472 152960688 SLC6A8 chrX 152990695 152991631 ABCD1 chrX 152994655 152994919 ABCD1 chrX 153001561 153001969 ABCD1 chrX 153002558 153002726 ABCD1 chrX 153005487 153005751 ABCD1 chrX 153005969 153006233 ABCD1 chrX 153008387 153008579 ABCD1 chrX 153008613 153008853 ABCD1 chrX 153008890 153009280 ABCD1 chrX 153215730 153215946 HCFC1 chrX 153216719 153216935 HCFC1 chrX 153217396 153217612 HCFC1 chrX 153223958 153224102 HCFC1 chrX 153228606 153228822 HCFC1 chrX 153229626 153229842 HCFC1 chrX 153230046 153230262 HCFC1 chrX 153237153 153237369 HCFC1 chrX 153640371 153640611 TAZ chrX 153641493 153641957 TAZ chrX 153642420 153642564 TAZ chrX 153647827 153648139 TAZ chrX 153648318 153648486 TAZ chrX 153648493 153648661 TAZ chrX 153648938 153649130 TAZ chrX 153649192 153649384 TAZ chrX 153760190 153760738 G6PD chrX 153760782 153761070 G6PD chrX 153761151 153761391 G6PD chrX 153761739 153761907 G6PD chrX 153762285 153762429 G6PD chrX 153762503 153762767 G6PD chrX 153763369 153763537 G6PD chrX 153764122 153764266 G6PD chrX 153774155 153774371 G6PD

Claims

The Claims We claim:

1. A method for multiplexed screening of subject samples for two or more target conditions, the method comprising:

(a) dividing a set of samples comprising nucleic acid markers for at least two target conditions for analysis into at least two sets of pools comprising a first set of X pools and a different second set of Y pools, wherein a subsample from every sample in the set of samples is included in the at least two sets of pools, wherein:

(i) X is the number of pools into which the samples are divided for the first set of pools; and

(ii) Y is the number of pools into which the samples are divided for the second set of pools; and

(iii) X and Y establish a pool matrix in which every sample in the set of samples has an intersection between the X pools and the Y pools; and

(b) coupling a nucleic acid barcode to the nucleic acid markers of each pool to yield barcoded nucleic acid markers, wherein the barcode is unique to the pool and differentiated from all other pools in the at least two sets of pools;

(c) sequencing the nucleic acid markers from the sets of pools, wherein each sample is sequenced at least once in the X pools and at least once in the Y pools;

(d) using a computer, deconvoluting the sequencing data by identifying the markers at the intersections of the pool matrix for which the sequencing data is indicative of the target condition.

2. The method of claim 1 wherein step (c) comprises: (a) pooling the sets of pools into a set of one or more master pools; and

(b) sequencing the nucleic acid markers from the set of master pools to produce the sequencing data.

3. The method of claim 2 wherein pooling the sets of pools into the set of one or more master pools produces one master pool.

4. The method of claim 2 wherein pooling the sets of pools into the set of one or more master pools produces two master pools.

5. The method of claim 2 wherein pooling the two sets of pools into the set of one or more master pools produces three master pools.

6. The method of claim 1 wherein X is greater than 12.

7. The method of claim 1 wherein X and Y are each greater than 12.

8. The method of claim 1 wherein X is greater than 20.

9. The method of claim 1 wherein X is greater than 20 and Y is greater than 12.

10. The method of claim 1 wherein X and Y are each greater than 20.

11. The method of claim 1 wherein X is greater than 50 and Y is greater than 12.

12. The method of claim 1 wherein X and Y are each greater than 50.

13. The method of any of claims 1 and following wherein the set of samples originate from a child.

14. The method of any of claims 1 and following wherein the set of samples originate from an infant.

15. The method of any claims 1 and following wherein the set of samples originate from a newborn infant.

16. The method of any claims 1 and following wherein the set of samples comprise nucleic acid markers for at least ten target conditions.

17. The method of any claims 1 and following wherein the set of samples comprise nucleic acid markers for at least 50 target conditions.

18. The method of any claims 1 and following wherein the set of samples comprise nucleic acid markers for at least 100 target conditions.

19. The method of any of claims 16-18 wherein the target conditions comprise the conditions set forth in Table 1.

20. The method of any of claims 16-18 wherein the target conditions comprise conditions selected from Table 2.

21. The method of any of claims 16-18 wherein the target conditions comprise conditions selected from Table 3.

22. The method of any of claims 16-18 wherein the target conditions comprise at least 10 conditions selected from Table 2.

23. The method of any of claims 16-18 wherein the target conditions comprise at least 10 conditions selected from Table 3.

24. The method of any of claims 16-18 wherein the target conditions comprise at least 20 conditions selected from Table 2.

25. The method of any of claims 16-18 wherein the target conditions comprise at least 20 conditions selected from Table 3.

26. The method of claim 1 further comprising referencing the sequencing data against a reference database of mutations and associated probabilities of conditions, to identify a probability that the sequencing data is indicative of the target condition.

27. The method of claim 26 wherein the database is maintained as part of a system in which the identification of the markers indicative of the target condition is correlated with subsequent empirical data from a sample and a probability of the target condition based on the empirical data is updated in the database.

28. The method of any of claims 1 and following further comprising using the computer to provide an output indicating when the target condition is not distinguishable between positive and carrier status.

29. The method of claim 28 wherein the computer output includes a recommendation for subsequent testing to make a definitive determination of positive or carrier status.

30. The method of claim 28 further comprising carrying out the subsequent testing, wherein subsequent testing comprises sequencing a nucleic acid sample from a subject.

31. The method of any of claims 1 and following further comprising using the computer to provide an output indicating when the deconvoluting of the sequencing data does not clearly identify individual samples from a master pool for which the sequencing data is indicative of the target condition.

32. The method of claim 31 wherein the computer recommends subsequent testing to make a definitive determination of an individual sample for which the sequencing data is indicative of the target condition.

33. The method of claim 31 further comprising carrying out the subsequent testing, wherein the subsequent testing comprises sequencing the nucleic acid sample from one or more subjects from the master pool from which the sequencing data does not clearly identify individual samples.

34. The method of any of claims 1 and following wherein the nucleic acid markers for analysis and the at least two target conditions are selected to result in a successful deconvolution rate of at least 50%.

35. The method of any of claims 1 and following wherein the nucleic acid markers for analysis and the at least two target conditions are selected to result in a successful deconvolution rate of at least 70%.

36. The method of any of claims 1 and following wherein the nucleic acid markers for analysis and the at least two target conditions are selected to result in a successful deconvolution rate of at least 95%.

37. The method of any of claims 1 and following wherein the samples are from a testing population and the target conditions comprise rare conditions which have an incidence rate in the testing population of less than 1 in 50.

38. The method of any of claims 1 and following wherein the samples are from a testing population and the target conditions comprise rare conditions which have an incidence rate in the testing population of less than 1 in 200.

39. The method of any of claims 1 and following wherein the samples are from a testing population and the target conditions comprise rare conditions which have an incidence rate in the testing population of less than 1 in 500.

40. The method of any of claims 1 and following wherein the samples are from a testing population and at least 50% of the target conditions have an incidence rate in the testing population of less than 1 in 50.

41. The method of any of claims 1 and following wherein the samples are from a testing population and at least 75% of the target conditions have an incidence rate in the testing population of less than 1 in 200.

42. The method of any of claims 1 and following wherein the samples are from a testing population and at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 500.

43. The method of any of claims 1 and following wherein the samples are from a testing population and X and Y are each at least 10 and at least 50% of the target conditions have the incidence rate in the testing population of less than 1 in 50.

44. The method of any of claims 1 and following wherein the samples are from a testing population and X and Y are each at least 50 and at least 75% of the target conditions have the incidence rate in the testing population of less than 1 in 200.

45. The method of any of claims 1 and following wherein the samples are from a testing population and X and Y are each at least 200 and at least 90% of the target conditions have the incidence rate in the testing population of less than 1 in 500.

46. The method of any of claims l and following wherein:

(a) the target conditions comprise at least 10 conditions,

(b) X and Y are each greater than 10; and

(c) at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 50.

47. The method of any of claims l and following wherein:

(a) the target conditions comprise at least 50 conditions,

(b) the sets of pools comprise samples from at least 50 subjects; and (c) at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 200.

48. The method of any of claims l and following wherein:

(a) the target conditions comprise at least 200 conditions;

(b) X and Y are each greater than 200; and

(c) at least 90% of the target conditions have an incidence rate in the testing population of less than 1 in 500.

49. The method of any of claims 1 and following wherein some or all of the markers are selected to give a yes/no answer with respect to one or more of the target conditions.

50. The method of any of claims 1 and following wherein some or all of the markers are selected to give a probability of a yes or a probability of a no with respect to one or more of the target conditions.

51. The method of any of claims 1 and following wherein the sample comprises nucleic acid markers obtained from blood, plasma, serum, urine, or another bodily fluid.

52. The method of any of claims 1 and following wherein the sample comprises nucleic acid markers obtained from a reconstituted blood spot.

53. The method of any of claims 1 and following wherein the target conditions comprise a disease for which newborns are screened.

54. The method of any of claims 1 and following wherein the target conditions are all diseases for which newborns are screened.

55. The method of any of claims 1 and following wherein the target conditions comprise an enzyme deficiency or an enzyme activity deficiency.

56. The method of any of claims 1 and following wherein the target conditions comprise a metabolic disorder.