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The efficacy of detecting variants with small effects on the Affymetrix 6.0 platform using pooled DNA.

ABSTRACT: Genome-wide genotyping of a cohort using pools rather than individual samples has long been proposed as a cost-saving alternative for performing genome-wide association (GWA) studies. However, successful disease gene mapping using pooled genotyping has thus far been limited to detecting common variants with large effect sizes, which tend not to exist for many complex common diseases or traits. Therefore, for DNA pooling to be a viable strategy for conducting GWA studies, it is important to determine whether commonly used genome-wide SNP array platforms such as the Affymetrix 6.0 array can reliably detect common variants of small effect sizes using pooled DNA. Taking obesity and age at menarche as examples of human complex traits, we assessed the feasibility of genome-wide genotyping of pooled DNA as a single-stage design for phenotype association. By individually genotyping the top associations identified by pooling, we obtained a 14- to 16-fold enrichment of SNPs nominally associated with the phenotype, but we likely missed the top true associations. In addition, we assessed whether genotyping pooled DNA can serve as an inexpensive screen as the second stage of a multi-stage design with a large number of samples by comparing the most cost-effective 3-stage designs with 80% power to detect common variants with genotypic relative risk of 1.1, with and without pooling. Given the current state of the specific technology we employed and the associated genotyping costs, we showed through simulation that a design involving pooling would be 1.07 times more expensive than a design without pooling. Thus, while a significant amount of information exists within the data from pooled DNA, our analysis does not support genotyping pooled DNA as a means to efficiently identify common variants contributing small effects to phenotypes of interest. While our conclusions were based on the specific technology and study design we employed, the approach presented here will be useful for evaluating the utility of other or future genome-wide genotyping platforms in pooled DNA studies.

SUBMITTER: Chiang CW

PROVIDER: S-EPMC3474315 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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The efficacy of detecting variants with small effects on the Affymetrix 6.0 platform using pooled DNA.

Chiang Charleston W K CW Gajdos Zofia K Z ZK Korn Joshua M JM Butler Johannah L JL Hackett Rachel R Guiducci Candace C Nguyen Thutrang T TT Wilks Rainford R Forrester Terrence T Henderson Katherine D KD Le Marchand Loic L Henderson Brian E BE Haiman Christopher A CA Cooper Richard S RS Lyon Helen N HN Zhu Xiaofeng X McKenzie Colin A CA Palmert Mark R MR Hirschhorn Joel N JN

Human genetics 20110322 5

Genome-wide genotyping of a cohort using pools rather than individual samples has long been proposed as a cost-saving alternative for performing genome-wide association (GWA) studies. However, successful disease gene mapping using pooled genotyping has thus far been limited to detecting common variants with large effect sizes, which tend not to exist for many complex common diseases or traits. Therefore, for DNA pooling to be a viable strategy for conducting GWA studies, it is important to deter ...[more]

PMID: 21424828

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Project description:BACKGROUND: Genome-wide association studies (GWAS) using array-based genotyping technology are widely used to identify genetic loci associated with complex diseases or other phenotypes. The costs of GWAS projects based on individual genotyping are still comparatively high and increase with the size of study populations. Genotyping using pooled DNA samples, as also being referred as to allelotyping approach, offers an alternative at affordable costs. In the present study, data from 100 DNA samples individually genotyped with the Affymetrix Genome-Wide Human SNP Array 6.0 were used to estimate the error of the pooling approach by comparing the results with those obtained using the same array type but DNA pools each composed of 50 of the same samples. Newly developed and established methods for signal intensity correction were applied. Furthermore, the relative allele intensity signals (RAS) obtained by allelotyping were compared to the corresponding values derived from individual genotyping. Similarly, differences in RAS values between pools were determined and compared. RESULTS: Regardless of the intensity correction method applied, the pooling-specific error of the pool intensity values was larger for single pools than for the comparison of the intensity values of two pools, which reflects the scenario of a case-control study. Using 50 pooled samples and analyzing 10,000 SNPs with a minor allele frequency of >1% and applying the best correction method for the corresponding type of comparison, the 90% quantile (median) of the pooling-specific absolute error of the RAS values for single sub-pools and the SNP-specific difference in allele frequency comparing two pools was 0.064 (0.026) and 0.056 (0.021), respectively. CONCLUSIONS: Correction of the RAS values reduced the error of the RAS values when analyzing single pool intensities. We developed a new correction method with high accuracy but low computational costs. Correction of RAS, however, only marginally reduced the error of true differences between two sample groups and those obtained by allelotyping. Exclusion of SNPs with a minor allele frequency of ? 1% notably reduced the pooling-specific error. Our findings allow for improving the estimation of the pooling-specific error and may help in designing allelotyping studies using the Affymetrix Genome-Wide Human SNP Array 6.0.

Project description:BackgroundCopy number data are routinely being extracted from genome-wide association study chips using a variety of software. We empirically evaluated and compared four freely-available software packages designed for Affymetrix SNP chips to estimate copy number: Affymetrix Power Tools (APT), Aroma.Affymetrix, PennCNV and CRLMM. Our evaluation used 1,418 GENOA samples that were genotyped on the Affymetrix Genome-Wide Human SNP Array 6.0. We compared bias and variance in the locus-level copy number data, the concordance amongst regions of copy number gains/deletions and the false-positive rate amongst deleted segments.ResultsAPT had median locus-level copy numbers closest to a value of two, whereas PennCNV and Aroma.Affymetrix had the smallest variability associated with the median copy number. Of those evaluated, only PennCNV provides copy number specific quality-control metrics and identified 136 poor CNV samples. Regions of copy number variation (CNV) were detected using the hidden Markov models provided within PennCNV and CRLMM/VanillaIce. PennCNV detected more CNVs than CRLMM/VanillaIce; the median number of CNVs detected per sample was 39 and 30, respectively. PennCNV detected most of the regions that CRLMM/VanillaIce did as well as additional CNV regions. The median concordance between PennCNV and CRLMM/VanillaIce was 47.9% for duplications and 51.5% for deletions. The estimated false-positive rate associated with deletions was similar for PennCNV and CRLMM/VanillaIce.ConclusionsIf the objective is to perform statistical tests on the locus-level copy number data, our empirical results suggest that PennCNV or Aroma.Affymetrix is optimal. If the objective is to perform statistical tests on the summarized segmented data then PennCNV would be preferred over CRLMM/VanillaIce. Specifically, PennCNV allows the analyst to estimate locus-level copy number, perform segmentation and evaluate CNV-specific quality-control metrics within a single software package. PennCNV has relatively small bias, small variability and detects more regions while maintaining a similar estimated false-positive rate as CRLMM/VanillaIce. More generally, we advocate that software developers need to provide guidance with respect to evaluating and choosing optimal settings in order to obtain optimal results for an individual dataset. Until such guidance exists, we recommend trying multiple algorithms, evaluating concordance/discordance and subsequently consider the union of regions for downstream association tests.

Dataset Information

The efficacy of detecting variants with small effects on the Affymetrix 6.0 platform using pooled DNA.

Publications

The efficacy of detecting variants with small effects on the Affymetrix 6.0 platform using pooled DNA.

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