[40], and has been successfully used for gene mapping in complex diseases [41–44]. Similar methods were used to identify the BRCA1 breast cancer susceptibility gene, where evidence for linkage was only present in earlier-onset families [22,45]. Subsets of families with earlier ages-of-onset may help evaluate genetic heterogeneity and define subgroups with a stronger genetic effect. A priori specification of the age-of-onset threshold is unnecessary, as OSA uses maximal linkage evidence to define subsets. OSA was conducted using nonparametric multipoint family-specific LODs from microsatellite genotypes from the original screen as input [7], with age-of-onset as a covariate. A permutation procedure provides empirical p-values for the significance of the increase in the maximum-subset-LOD from the overall LOD. To assess SNP linkage, two-point LODs were calculated using Merlin [46]. The Pedigree Disequilibrium Test (PDT) and Genotype-PDT were used for family-based association. PDT, an extension of the transmission-disequilibrium-test (TDT), allows incorporation of extended pedigrees and is valid even with population substructure [47]. Power calculations (QUANTO:http://hydra.usc.edu/) showed power $0.80 to detect effect sizes of $2.45 for the lowest allele frequency SNP (0.02, rs16139), and effect sizes $1.35 for the highest allele frequency SNP (0.49, rs16147). In CATHGEN, association was assessed using logistic regres- sion models adjusting for race and sex; and for race, sex, hypertension, diabetes, dyslipidemia, smoking and body-mass- index (BMI) (multivariable model). Measured genotype analysis using generalized linear models was performed to compare differences in means of quantitative traits (NPY levels, age-of- onset) by NPY genotype. Baseline differences were assessed using a chi-square or t-test. The Graphical Overview of Linkage Disequilibrium (GOLD) program [48] was used to assess LD. Haplotype analysis used HaploStats 1.1.0 (Mayo Clinic, Roche- ster, MN). We report p-values uncorrected for multiple compar- isons, but also present results in the context of correction for LD between SNPs [35]. The extent of atherosclerosis was compared between control and NPY1 receptor antagonist groups in mice with one-way ANOVA and Tukey’s post-hoc test for multiple References 1. American Heart Association (2006) Heart and Stroke Statistical Update. http:// www.americanheart.org/statistics. 2. Marenberg ME, Risch N, Berkman LF, Floderus B, De Faire U (1994) Genetic susceptibility to death from coronary heart disease in a study of twins. N Engl J Med 330: 1041–1046. 3. Rissanen AM (1979) Familial occurrence of coronary artery diease: effect of age at diagnosis. Am J Cardiol 44: 60–66. 4. Connelly JJ, Wang T, Cox JE, Haynes C, Wang L, et al. (2006) GATA2 Is Associated with Familial Early-Onset Coronary Artery Disease. PLoS Genet 2. 5. 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Hall JM, Lee MK, Newman B, Morrow JE, Anderson LA, et al. (1990) Linkage of early-onset familial breast cancer to chromosome 17q21. Science 250: 1684–1689. PLoS Genetics | www.plosgenetics.org 9 January 2009 | Volume 5 | Issue 1 | e1000318