March 2009
Volume 50, Issue 3
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Cornea  |   March 2009
Linkage Analysis in Keratoconus: Replication of Locus 5q21.2 and Identification of Other Suggestive Loci
Author Affiliations
  • Luigi Bisceglia
    From the Medical Genetics Service, the
  • Patrizia De Bonis
    From the Medical Genetics Service, the
  • Costantina Pizzicoli
    Institute of Ophthalmology, University of Foggia, Foggia, Italy.
  • Lucia Fischetti
    From the Medical Genetics Service, the
  • Antonio Laborante
    Department of Ophthalmology, and the
  • Michele Di Perna
    From the Medical Genetics Service, the
  • Francesco Giuliani
    Scientific Direction, IRCCS-CSS Hospital, San Giovanni Rotondo, Italy; and the
  • Nicola Delle Noci
    Institute of Ophthalmology, University of Foggia, Foggia, Italy.
  • Luca Buzzonetti
    Department of Ophthalmology, and the
  • Leopoldo Zelante
    From the Medical Genetics Service, the
Investigative Ophthalmology & Visual Science March 2009, Vol.50, 1081-1086. doi:10.1167/iovs.08-2382
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      Luigi Bisceglia, Patrizia De Bonis, Costantina Pizzicoli, Lucia Fischetti, Antonio Laborante, Michele Di Perna, Francesco Giuliani, Nicola Delle Noci, Luca Buzzonetti, Leopoldo Zelante; Linkage Analysis in Keratoconus: Replication of Locus 5q21.2 and Identification of Other Suggestive Loci. Invest. Ophthalmol. Vis. Sci. 2009;50(3):1081-1086. doi: 10.1167/iovs.08-2382.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

purpose. Keratoconus (KC) is the most common indication for corneal transplantation in the Western world, with etiologic mechanisms still poorly understood. The disease prevalence in the general population is approximately 1:2000, and familial aggregation, together with increased familial risk, suggests important genetic influences on its pathogenesis. To date, several loci for familial keratoconus have been described, without the identification of any responsible gene in the respective mapped intervals. The aim of this study was to identify causative/susceptibility genes for keratoconus.

methods. A total of 133 individuals (77 affected and 59 unaffected) of 25 families from southern Italy were genotyped using microsatellite markers and included in a genome-wide scan. Nonparametric and parametric analysis using an affected-only strategy were calculated by using genetic algorithm software.

results. The chromosomal regions 5q32-q33, 5q21.2, 14q11.2, 15q2.32 exhibited the strongest evidence of linkage by nonparametric analysis (NPL = 3.22, 2.73, 2.62, and 2.32, respectively). The regions 5q32-q33 and 14q11.2 were also supported by multipoint parametric analysis, for which heterogeneity LOD (HLOD) scores of 2.45 (α = 0.54) and 2.09 (α = 0.46), respectively, were obtained under an affected-only dominant model.

conclusions. This study represents the first KC linkage replication study on the chromosomal region 5q21.2 and reports evidence of suggestive linkage in several regions for which suggestive or significant linkage has been previously detected in different populations.

Keratoconus (KC; Mendelian Inheritance in Man [MIM] 148300) is a degenerative noninflammatory disease of the cornea, progressive in 20% of cases, characterized by changes in corneal collagen structure, 1 2 organization, 3 and intercellular matrix. 4 Additional characteristics include apoptosis 5 and necrosis of keratinocytes 6 involving the central anterior stroma and the Bowman’s lamina, 7 which produces the typical weakening of the corneal tissue. It is a major indication for lamellar or perforating keratoplasty in the Western world, 8 9 with a prevalence in the general population approximately of 1:2000. 10 11 The disease arises in the teenage years with progressive myopia and astigmatism; usually, it arrests in the third or fourth decade of life. 
KC can be diagnosed by well-recognized clinical signs, including stromal thinning, Vogt’s striae, Fleischer’s ring, and scissoring of the retinoscopic reflex with a fully dilated pupil. The most sensitive and accurate diagnostic method is the computer-assisted videokeratography, which allows detection of KC at an early stage, before clinical signs are evident. 12 13 Histologic observations have demonstrated degradation of the corneal epithelium basal membrane, diminution of the number of collagen fibrils, thinning of the corneal stroma, and keratocyte apoptosis. 4 6  
Biochemical studies describe increased activity of metalloproteinase (MMP-2 and -9) and lower expression of protease inhibitors such as α1-protease inhibitor. 14 15  
Even if the etiology and the mechanisms leading to the KC are still unknown, there is evidence suggesting an important role of genetic factors in determinism of the disease. In nearly all patients, KC is an isolated defect; however, in some cases it may be a symptom in syndromic conditions, as observed in Ehlers-Danlos, Marfan, Apert, Noonan, and Down syndromes. KC prevalence in first-degree relatives of KC patients is 3.34%, 15 to 67 times higher than in general population. 11 Familial transmission have been demonstrated in 6% to 23.5% of cases described in literature, 11 16 17 18 19 and concordance is high among monozygotic twins. 20  
Both recessive and dominant patterns of inheritance have been described. Autosomal dominant inheritance has more frequently been reported in families, showing incomplete penetrance and variable expressivity. Subtle videokeratographic anomalies have been reported among relatives of KC patients, allowing the detection of low expressivity forms of KC, usually referred as “subclinical” or “forme fruste”. 18 21 22 Multifactorial inheritance and a major gene model have also been proposed. 16 23 24  
In the past few years, several authors searched to identify KC causative/susceptibility genes by applying candidate gene and linkage analysis. To date, candidate loci for KC have been mapped by genome wide scan to the chromosomal regions 2p24, 3p14-q13, 5q14.3-q21.1, and 16q22.3-q23.1, 25 26 27 28 29 in large single or multiple pedigree showing an autosomal dominant model of transmission; and to the chromosomal regions 4q31, 5q31, 9q34, 12p12, 14p11, 17q24, and 20q12, 24 30 by using an affected-only linkage analysis that implies a multifactorial model of transmission (Table 1) . Several candidate genes, COL6A1, SOD1, MMP9, MMP2, COL8A1 located on candidate loci were excluded as causative genes 26 29 30 31 32 while mutations in the visual system homeobox gene (VSX1) has been reported in a small number of patients affected by KC. 19 33 34  
The evidence of KC linked to multiple chromosomal regions is consistent with both an oligo/polygenic and a multiple-susceptibility gene model, all having a small or moderate effect in determining the pathologic phenotype, as observed in complex diseases. 
The aim of this study was to identify causative/susceptibility genes for KC by analyzing a series of 25 families from Southern Italy. 
Methods
Families
A total of 207 unrelated individuals with KC from southern Italy were recruited at the Medical Genetic Service of IRCCS Hospital Casa Sollievo della Sofferenza (San Giovanni Rotondo, Italy). Of these, 54 patients (26%) were familial cases. The analyzed population in the present study is comprised of 25 pedigrees including 133 subjects, of which 77 are affected and 59 unaffected. 
Two affected individuals were present in 10 pedigrees, 3 in 8 pedigrees, 4 in 2 pedigrees, and 5 in 5 pedigrees. On the whole, studied subjects resulted in 42 affected sibling pairs, 3 affected grandparent-grandchild pairs, 17 affected uncle-nephew pairs, and 10 affected first cousins pairs. The diagnosis of KC was made on the basis of history of penetrating keratoplasty, clinical examination (corneal stromal thinning, Vogt’s striae, Fleischer’s ring, Munson’s sign), and videokeratographic evaluation. The KISA% index was used to quantify the irregular corneal shape and astigmatism to assess KC according to previously published criteria. 18 19 35 The local Ethics Committee at IRCCS Hospital Casa Sollievo della Sofferenza approved the study. Venous blood was collected from the patients and family members after informed consent. The research adhered to the tenets of the Declaration of Helsinki. 
Genome-wide Scan
DNA was extracted from peripheral lymphocytes by standard phenol-chloroform methodology. A genome-wide scan was performed by genotyping 382 highly polymorphic markers positioned along the 22 autosomal chromosomes at ∼10 cM density included in the linkage mapping panel (ABI PRISM Linkage Mapping Set version 2.5-MD10; Applied Biosystems, Foster City, CA). Refinement of the interesting chromosomal regions identified was performed by analyzing additional microsatellite markers at approximately 2 cM density according to available maps: Marshfield genetic map (http://research.marshfieldclinic.org/genetics/GeneticResearch/compMaps.asp) and University of California Santa Cruz draft of the human genome (www.genome.ucsc.edu). PCR reactions were performed in a 15 μL reaction volume containing 60 ng of genomic DNA; 5 pmoles of each primer (one of which was fluorescently labeled); PCR buffer including 50 mM KCl, 10 mM Tris-HCl (pH 8.3), 1.5 mM MgCl2, and 200 μM each of dATP, dGTP, dTTP, and dCTP; and 1 U of DNA polymerase (AmpliTaq; Applied Biosystems). Reaction mixtures were heated to 94°C for 10 minutes and then cycled 30 times as follows: 30 seconds at 94°C, 30 seconds at 55°C, and 30 seconds at 72°C and a final elongation step at 72°C for 7 minutes. The fluorescence-labeled PCR products were pooled (10 to 20 markers/pool). Two microliters of the pooled products were mixed with 0.3 μL of the internal size standard and 10 μL of deionized formamide, denatured, and separated using a capillary electrophoresis apparatus (ABI PRISM 3100 DNA sequencer; Applied Biosystems). The fragment analysis was carried out by using genotyping software (GeneScan 3.7 and Genotyper 3.7 NT; both from Applied Biosystems). 
The inheritance of each marker in all pedigrees was tested by software for identifying inconsistencies (PedCheck) due to null alleles, mistyping, no paternity, or other errors, and repeat genotyping was performed as necessary. 36 All marker alleles were considered to have equal frequency. 
Statistical Analysis
Multipoint nonparametric LOD score (NPL all statistic) and parametric heterogeneity LOD score (HLOD), with the assumption of an autosomal dominant mode of inheritance, with disease allele frequency of 0.001 were calculated using statistical genetic analysis software (Genehunter). 37 Parametric HLOD scores were evaluated using an affected-only strategy that is, by assuming all unaffected individuals as phenotypically unknown, since they may not provide reliable information on the underlying disease-locus genotype for a complex disease. Significant and suggestive linkage was defined according to the criteria proposed by Lander and Kruglyak 38 who stated that: i) for studies involving a mixture of relative types, by using allele-sharing methods in affected relative pairs, the thresholds of P value are in the range of 10−3 to 5 × 10−4 (LOD = 1.9 to 2.4) for suggestive linkages and 5 × 10−5 to 10−5 (LOD = 3.3 to 3.8) for significant linkages; and ii) the term of replication study should be reserved for situations in which significant linkage has already been obtained in an initial study. In this case, a P value of 0.01 should be required to declare confirmation at the 5% level. 
Results
Genome-wide Analysis
We undertook a genome-wide scan for linkage in 133 individuals of 25 families; results are represented in Figure 1 . Nonparametric and parametric analysis using an affected-only strategy found several regions suggestive of linkage at the genome-wide level. Nonparametric analysis showed strongest evidence of nearly significant linkage at the genome-wide level on the region 5q32-q33, yielding a NPL=3.33 (P = 0.000559) and of suggestive linkage on the regions 5q21.2 (NPL = 2.48, P = 0.007091), 9q21.13 (NPL = 2.54, P = 0.005965), and 14q11.2 (NPL = 2.93, P = 0.002013). In addition, on the regions 5q32-q33 and 14q11.2, the maximum HLOD score was suggestive of linkage, 2.68 (α = 0.64) and 2.91 (α = 0.58), respectively. Other regions showing suggestive linkage were identified on chromosomes 1q, 2p, 10q, 15q, 16q, and 18p. 
Fine Mapping
A total of additional 67 microsatellite markers on chromosomes 1, 2, 5, 9, 10, 14, 15, 16, and 18, along the previous identified regions were genotyped to increase the resolution map and multipoint nonparametric and parametric linkage analysis was performed. After the analysis, all peak regions (with exception of those on chromosomes 1, 2, and 10) showed evidence of suggestive linkage for at least one of the statistics used to analyze our data set of KC families. In particular, on the region 5q32-q33 the NPL peak slightly decreased to 3.22 (P = 0.000836) remaining, nevertheless, highly significant at level of suggestive linkage. A slight decrease of NPL peak was also observed for the regions 9q21.13, 14q11.2, and 16q23.1, whereas a slight increase of the peak was obtained for the regions 5q21.2, 9q22.2, 15q15.1, and 18p11.13. Results of parametric and nonparametric analysis after fine mapping are summarized in Table 2and Figure 2
Discussion
A genome-wide scan was conducted to identify causative/susceptibility genes for KC in a series of pedigrees by applying nonparametric and parametric affected-only analysis, which are more useful statistic methods to detect loci for complex disease. 
Evidence of linkage was observed on several chromosomal loci, most of which were confirmed after fine mapping. The linkage peaks obtained satisfy criteria for suggestive linkage, according to those proposed by Lander and Kruglyak. 38 The more relevant results were obtained for the regions 5q32-q33, 5q21.2, 14q11.2, and 15q15.1 (Table 2) . Interestingly, the comparison of our results with those obtained by other authors in different populations 24 25 28 showed that some peak regions were partially or completely overlapping. 
On the long arm of chromosome 5, we identified a large region spanning approximately 70 cM, in which NPL analysis was significant at a P value ≤ 0.05. At the boundaries of this region we observed two intervals, 109 to 135 cM and 153 to 171 cM, respectively, in which the P value reached a level of significance ≤ 0.01. The higher peaks of suggestive linkage were found at 116 cM and 167 cM, respectively. 
The first one falls in 5q21.2 (NPL = 2.73, P = 0.003669) overlapping with the region (positioned between 100 to 120 cM) reported by Tang et al. 28 (LOD = 3.53) in a large four-generation pedigree. Thus, very likely, the two linkage regions, overlapping on the interval between 109 to 120 cM, identify the same putative locus for KC. Our results provide the first evidence of linkage replication on the 5q21.2 region, in accordance with the findings of Lander and Kruglyak. 38 Our second linkage peak yielded a nearly significant linkage in 5q32-q33 (NPL = 3.22, P = 0.000836) overlapping with the region identified by Li et al. 24 at position 157 cM (NPL=2.03) in Caucasian individuals, and at position 143 cM (NPL = 2.90), in Caucasian and Hispanic individuals. Moreover, we found a suggestive linkage (NPL = 2.62, P = 0.004994) in 14q11.2, the same region in which Li et al. reported a suggestive linkage in Caucasian persons (NPL = 2.91). For both 5q32-q33 and 14q11.2 regions, the linkage did not reach a significant level in our study or the study of Li et al. 24 Nonetheless, this highly suggestive linkage, independently found in two different data sets of families, provides further evidence that these could be loci associated with KC development. 
Other regions showing suggestive linkage were also found on chromosomes 9q21-q22 (60 and 82 cM), 15q15 (38 cM), 18p11 (17 cM), and 16q23 (106 cM), the last lying close to the 16q22.3-q23.1 region, identified by Tyynismaa et al. 25  
Several hundreds of known and predicted genes are mapped in the regions identified in our study, but none of these can be considered obvious candidates. The following genes, LOX (5q23.2), SPARC (5q31.3-q32), PCSK5 (9q21.3), CTSL (9q21-q22), ANXA1 (9q12-q21.2), APEX1 (14q11.2-q12), and CDK10 (16q24), could be considered as possible KC candidate genes on the basis of their known function (proteinase, matrix-associated proteins, cellular proliferation) and/or their KC corneal expression. 
LOX encodes for lysil oxidase, an extra cellular matrix protein enzyme that initiates the cross-linking of collagens and elastin. These gene was found upregulated (signal ratio of 1.5) in KC epithelium 39 and was indicated as candidate gene because of its position. 24 SPARC encodes for a matrix-associated protein (secreted protein acidic and rich in cysteine/osteonectin/BM40) that elicits changes in cell shape, inhibits cell-cycle progression, and influences the synthesis of extracellular matrix (ECM). 40 Sparc-deficient mice were observed normal and fertile until 6 months of age, when they developed a severe eye pathology characterized by cataract formation and rupture of the lens capsule. 41 APEX1 gene encodes for APEX nuclease (multifunctional DNA repair enzyme) 1, also called apurinic endonuclease (APE), a DNA repair enzyme involved in a caspase-independent cell death pathway. 42 PCSK5 encodes for a proprotein convertase that mediates posttranslational endoproteolytic processing for several integrin alpha subunits; in vascular smooth muscle cell, PCSK5 is necessary for endoproteolytic activation of integrin α-V, which leads to integrin-mediated cell adhesion, migration, and signaling. 43 CTSL encodes for cathepsin L, a lysosomal cysteine proteinase that plays a major role in intracellular protein catabolism. It also shows the most potent collagenolytic and elastinolytic activity in vitro of any of the cathepsins (OMIM: 116880). ANXA1 encodes for Annexin I belonging to a family of Ca(2+)-dependent phospholipid binding proteins, having a potential anti-inflammatory activity. It has been found upregulated (signal ratio of 1.5) in KC corneal epithelium. 39 CDK10 (PISSLRE) encodes a protein belonging to the CDK subfamily of the Ser/Thr protein kinase family. This kinase has been shown to play a role in cellular proliferation and its function is limited to cell cycle G2-M phase. 44  
Genetic studies performed on familial KC showed a high level of genetic heterogeneity, if KC is analyzed as a dominant Mendelian trait with incomplete penetrance and variable expressivity. In the same way, multiple susceptibility-disease loci were also identified considering KC as a complex trait. Both transmission models could explain the familial aggregation and the variability of clinical manifestation of KC (unilateral or bilateral disease, variable onset and progression, presence of subclinical or forme fruste). In addition, the genetic heterogeneity could suggest the hypothesis that mutations in several different genes, involved in related pathways, may act on common targets responsible for the disease. 
In conclusion, this study represents the first KC linkage replication study on the chromosomal region 5q21.2 and reports evidence of suggestive linkage in several regions for which suggestive or significant linkage has been previously detected in different populations. 
 
Table 1.
 
Chromosome Loci Reported to Show Evidence for Linkage with Keratoconus
Table 1.
 
Chromosome Loci Reported to Show Evidence for Linkage with Keratoconus
Study Design No. of Families No. of Individuals (No. of affected) LOD or HLOD (α) NPL (P) Chromosome Region (peak cM) Sample Examined References
Extended pedigrees (AD) 28 253 (112) 5.13 (α = 0.52) 3.26 (P < 0.005) 2p24 (40) Caucasian, Arab, Caribbean, African 27
1 21 (11) 3.09 3p14-q13 (108) Italian 26
1 9 (7) 3.53 5q14.3-q21.1 (103) Caucasian 28
1 30 (16) 8.13 15q22.33-24.2* (74) Irish 29
20 76 (42) 4.10 3.27 (P = 0.00006) 16q22.3-q23.1 (75) Finnish 25
Allele-sharing (IBD) (8) (P = 0.00021) 20q12 Northwest Tasmania 30
Affected sib pairs 17 93 (≥17 sib pairs) 2.3 0.96 (P = 0.008) 2q (184) Hispanic 24
17 93 (≥17 sib pairs) 2.2 1.69 (P = 0.006) 3p (74) Hispanic 24
67 351 (110 sib pairs) 2.2 2.68 (P = 0.001) 4q (176) Caucasian/Hispanic 24
67 351 (110 sib pairs) 2.01 2.90 (P = 0.013) 5q31 (143) Caucasian/Hispanic 24
17 93 (≥17 sib pairs) 2.5 2.64 (P = 0.002) 5p (42) Hispanic 24
17 93 (≥17 sib pairs) 3.8 3.55 (P = 0.001) 9p (34) Hispanic 24
67 351 (110 sib pairs) 3.5 2.83 (P < 0.001) 9q (160) Caucasian/Hispanic 24
40 217 (≥40 sib pairs) 2.3 1.98 (P = 0.003) 11p (12) Caucasian 24
67 351 (110 sib pairs) 2.5 2.64 (P = 0.010) 12p (7) Caucasian/Hispanic 24
67 351 (110 sib pairs) 2.6 2.23 (P = 0.032) 14p (19) Caucasian/Hispanic 24
17 93 (≥17 sib pairs) 3.9 3.32 (P < 0.001) 17q(86) Hispanic 24
Figure 1.
 
Multipoint linkage results of the genome-wide scan for keratoconus in 25 pedigrees except for chromosome X.
Figure 1.
 
Multipoint linkage results of the genome-wide scan for keratoconus in 25 pedigrees except for chromosome X.
Table 2.
 
Suggestive Linkage Peaks for Keratoconus after Fine Mapping
Table 2.
 
Suggestive Linkage Peaks for Keratoconus after Fine Mapping
Chromosome Region Position (cM) HLOD (α) NPL (P Value)
5q21.2 116 0.49 (0.23) 2.73 (0.003669)
5q32-q33 167 2.45 (0.54) 3.22 (0.000836)
9q21.13 60 1.07 (0.30) 1.93 (0.027194)
9q22.2 82 1.61 (0.40) 2.10 (0.018552)
14q11.2 1 2.09 (0.46) 2.62 (0.004994)
15q15.1 38 1.74 (0.42) 2.32 (0.010852)
16q23.1 106 0.45 (0.26) 1.97 (0.025172)
18p11.31 17 0.72 (0.27) 2.00 (0.023258)
Figure 2.
 
Linkage results of fine mapping regions showing suggestive evidence of linkage.
Figure 2.
 
Linkage results of fine mapping regions showing suggestive evidence of linkage.
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Figure 1.
 
Multipoint linkage results of the genome-wide scan for keratoconus in 25 pedigrees except for chromosome X.
Figure 1.
 
Multipoint linkage results of the genome-wide scan for keratoconus in 25 pedigrees except for chromosome X.
Figure 2.
 
Linkage results of fine mapping regions showing suggestive evidence of linkage.
Figure 2.
 
Linkage results of fine mapping regions showing suggestive evidence of linkage.
Table 1.
 
Chromosome Loci Reported to Show Evidence for Linkage with Keratoconus
Table 1.
 
Chromosome Loci Reported to Show Evidence for Linkage with Keratoconus
Study Design No. of Families No. of Individuals (No. of affected) LOD or HLOD (α) NPL (P) Chromosome Region (peak cM) Sample Examined References
Extended pedigrees (AD) 28 253 (112) 5.13 (α = 0.52) 3.26 (P < 0.005) 2p24 (40) Caucasian, Arab, Caribbean, African 27
1 21 (11) 3.09 3p14-q13 (108) Italian 26
1 9 (7) 3.53 5q14.3-q21.1 (103) Caucasian 28
1 30 (16) 8.13 15q22.33-24.2* (74) Irish 29
20 76 (42) 4.10 3.27 (P = 0.00006) 16q22.3-q23.1 (75) Finnish 25
Allele-sharing (IBD) (8) (P = 0.00021) 20q12 Northwest Tasmania 30
Affected sib pairs 17 93 (≥17 sib pairs) 2.3 0.96 (P = 0.008) 2q (184) Hispanic 24
17 93 (≥17 sib pairs) 2.2 1.69 (P = 0.006) 3p (74) Hispanic 24
67 351 (110 sib pairs) 2.2 2.68 (P = 0.001) 4q (176) Caucasian/Hispanic 24
67 351 (110 sib pairs) 2.01 2.90 (P = 0.013) 5q31 (143) Caucasian/Hispanic 24
17 93 (≥17 sib pairs) 2.5 2.64 (P = 0.002) 5p (42) Hispanic 24
17 93 (≥17 sib pairs) 3.8 3.55 (P = 0.001) 9p (34) Hispanic 24
67 351 (110 sib pairs) 3.5 2.83 (P < 0.001) 9q (160) Caucasian/Hispanic 24
40 217 (≥40 sib pairs) 2.3 1.98 (P = 0.003) 11p (12) Caucasian 24
67 351 (110 sib pairs) 2.5 2.64 (P = 0.010) 12p (7) Caucasian/Hispanic 24
67 351 (110 sib pairs) 2.6 2.23 (P = 0.032) 14p (19) Caucasian/Hispanic 24
17 93 (≥17 sib pairs) 3.9 3.32 (P < 0.001) 17q(86) Hispanic 24
Table 2.
 
Suggestive Linkage Peaks for Keratoconus after Fine Mapping
Table 2.
 
Suggestive Linkage Peaks for Keratoconus after Fine Mapping
Chromosome Region Position (cM) HLOD (α) NPL (P Value)
5q21.2 116 0.49 (0.23) 2.73 (0.003669)
5q32-q33 167 2.45 (0.54) 3.22 (0.000836)
9q21.13 60 1.07 (0.30) 1.93 (0.027194)
9q22.2 82 1.61 (0.40) 2.10 (0.018552)
14q11.2 1 2.09 (0.46) 2.62 (0.004994)
15q15.1 38 1.74 (0.42) 2.32 (0.010852)
16q23.1 106 0.45 (0.26) 1.97 (0.025172)
18p11.31 17 0.72 (0.27) 2.00 (0.023258)
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