©2011 McGregor et al; licensee BioMed Central Ltd.
Common polymorphisms in human lysyl oxidase genes are not associated with the adolescent idiopathic scoliosis phenotype
1Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232 USA
2Center for Human Genetics Research, Vanderbilt University School of Medicine, Nashville, TN 37232 USA
3Department of Neurology, Washington University School of Medicine, St. Louis, MO 63110 USA
4Department of Pediatrics, Washington University School of Medicine, St. Louis, MO 63110 USA
5Department of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110 USA
6St. Louis Shriners Hospital for Children, St. Louis, MO 63131 USA
7Sarah M. and Charles E. Seay Center for Musculoskeletal Research, Texas Scottish Rite Hospital for Children, Dallas, TX 75219 USA
8Department of Orthopaedic Surgery, University of Texas Southwestern Medical Center, Dallas, TX 75390 USA
9McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, TX 75390 USA
10Department of Orthopedic Surgery and Rehabilitation, University of Iowa, Iowa City, IA 52242 USA
11The Perinatal Research Center, Nashville, TN, 37203 USA
12Department of Epidemiology, Rollins School of Public Health, Emory University, Atlanta, GA, 30322 USA
13Vanderbilt Kennedy Center for Human Development, Vanderbilt University, Nashville, TN 37232 USA
14Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Vanderbilt University, Nashville, TN 37232 USA
Received March 14, 2011; Accepted July 8, 2011.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0
), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
For initial SNP selection, the Tagger algorithm initially identified a total of 132 candidate coding and tag SNPs. After reiterations to accommodate the Sequenom Genotyping platform, final selection included 112 SNPs. These were genotyped in 138 case and 411 control samples (Additional File 1
). Of these, 9 SNPs and 15 samples (8 cases and 7 controls) failed genotyping and were excluded from further analyses. A total of 16 SNPs in LOXL1
, and LOXL4
(Table ) showed a difference by genotype with an uncorrected adjusted p < 0.1. These SNPs were carried forward for genotyping in the replication population.
SNPs genotyped in discovery cases and controls with p <0.1
Of the 16 SNPs carried forward to the replication samples, rs751160 (LOXL4) and rs12442211 (LOXL1) were not successfully genotyped. In the cohort of 400 cases and 506 controls, 17 samples (7 cases and 10 controls) failed genotyping. A total of 14 SNPs were analyzed within the replication cohort using the same two degree of freedom genotypic model with multivariate logistic regression, adjusting for gender. The minor allele frequencies were consistent between populations (Figure ). No SNPs showed a difference by genotype between cases and controls with p < 0.004 (Table ). Because the cases in the replication cohort were obtained from two different recruiting centers, the analysis was stratified by recruiting center to assess for confounding. No SNP attained significance in a single cohort that was masked by combining the analysis. In addition, the analysis was stratified by gender to determine if gender was acting as an effect modifier. No significant associations or evidence for effect modification were detected.
Figure 1 Minor allele frequency comparison between populations. The minor allele frequencies of the SNPs successfully genotyped in the replication phase are represented. The minor allele frequency did not differ significantly between the six populations as assessed (more ...)
Significance and odds ratios of SNPs genotyped in the replication phase
A secondary analysis of traditional allele association was performed for the SNPs genotyped in the replication cohort. The results indicate that no SNPs were significantly correlated after accounting for multiple testing.
Retrospective power calculations with performed with measured allele frequencies. Our discovery cohort of 138 cases and 411 controls had allele frequencies ranging from 0.06 - 0.49. We had 80% power to detect minimum odds ratios in the range of 1.7 - 2.1 using a 2 degree of freedom genotypic test with alpha = 0.1 (cutoff for inclusion in replication set). Assumptions included genotyping of the causative SNP, a dominant model without additive or multiplicative effects, and a population prevalence of 3%. In the replication cohort of 400 cases and 506 controls, the measured allele frequencies ranged from 0.13 - 0.46. This allowed for 80% power to detect minimum odds ratios in the range of 1.5 - 1.6 with the same parameters. The detection limit increased to 2.6 - 3.6 with a Bonferroni correction and alpha = 0.00357. Note the lowest minor allele frequency measured in the discovery cohort was 0.06, with minimum detectable odds ratio of 2.0. No SNPs carried forward in the replication set had a measured allele frequency of less than 0.13, indicating that rare polymorphisms were not fully assessed in this study.
Additional polymorphisms are added to public databases such as dbSNP with each build. To assess for the extent of coverage, the successfully genotyped SNPs were tested as proxy markers for all SNPs with allele frequency > 0.1 within 100 kb of each lysyl oxidase gene included in the 1000 Genomes pilot data release. We achieved an average r2 of 0.76 over 131 SNPs in LOX, 0.68 over 404 SNPs in LOXL1, 0.60 over 741 SNPs in LOXL2, 0.36 over 224 SNPs in LOXL3, and 0.64 over 344 SNPs in LOXL4.