On-Line Supplementaldata

On-line SupplementalData

Appendix

Genotyping

APOE alleles (corresponding to allele combinations at SNP +3937/rs429358 and SNP +4075/rs7412) were genotyped as previously reported 1. Genotyping efficiency was greater than 90% and quality control (QC) was achieved by including two sets of 12 control samples, and four sets of two standard samples on each 384-well plate. The laboratory technicians were blinded to family relations, affection status, and quality control samples.[PIL1]

Family-based association tests

These two programs have their own unique strengths that complement each other. PDT compares allele frequencies in affected and unaffected sibs 2. APL evaluates associations in different types of families, such as affected sibling pair, discordant sibling pair, and affected sibling pair plus one unaffected sibling. Finally, we used the program APL to perform haplotype-based association tests. Again, we evaluated associations in the overall families, the earlier-onset subset, and the later-onset subset to examine AAO-dependent associations.

We chose the false discovery rate (FDR) procedure introduced by Benjamini and Hochberg 3 to perform multiple testing corrections. FDR is the expected rate of false rejections of the null hypothesis among all discoveries. To obtain the FDR, we ranked all of the markers by the original p-values from smallest to largest, and then assessed the significance of each p-value based on their corresponding cutoff values. The cutoff is Pi= 0.05 x i/m for the ith smallest p-value for a total of m observed p-values. P-values lower than the respective cutoffs were considered significant.

Multi-locus association tests

Since higher-order locus-locus interaction may lead to computational complexity and difficult interpretation of results, we limited our search to the best one-locus, two-locus, and three-locus models. We used the extended multi-factor dimensionality reduction (EMDR) program 4 to examine possible multi-locus effects in our unrelated case-control sample.

Reference List

1. Saunders AM, Strittmatter WJ, Schmechel D et al. Association of apolipoprotein E allele epsilon 4 with late-onset familial and sporadic Alzheimer's disease. Neurology 1993;43(8):1467-1472.

2. Martin ER, Monks SA, Warren LL, Kaplan NL. A test for linkage and association in general pedigrees: the pedigree disequilibrium test. Am J Hum Genet 2000;67(1):146-154.

3. Benjamini Y, Hochberg Y. Controlling the false discovery rate: A practical and powerful approach to multiple testing. Journal of Royal Statistical Society 1995;57:289-300.

4. Mei H, Ma DQ, Ashley-Koch A, Martin ER. Extension of multifactor dimensionality reduction for identifying multi-locus effects in the GAW14 simulated data. BMC Genetics 2005;6(Suppl 1)(S53).

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