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Quality Management "Genotyping"

Quality management coordinator:
Dominik Seelow
Cologne Center for Genomics
Universität Köln
Phone: + 49 30 32639 210
E-Mail:
dominik.seelow@uni-koeln.de
 
It is one of the key missions of the National Genome Research Network (NGFN), to identify the genetic factors involved in complex diseases by establishing the link between phenotypic data and the genome of the patients. This approach requires substantial resources in high-throughput genotyping of “short tandem repeat” (STR) and “single nucleotide polymorphism” (SNP) markers. Therefore, a consortium was founded comprising the most potent genotyping centers of Germany to form a National Genotyping Platform (NGP).

Detection and elimination of genotyping errors is a major task, which has to precede data analysis. Genotype call rate (per marker, per sample, and globally), single genotype quality scores or confidence values (defined by the manufacturer, and averaged over markers and samples), sample allele frequency estimates, deviations from Hardy-Weinberg genotype proportions, heterozygote frequencies, and identity-by-state (IBS) values of samples constitute, among other quantities, a set of parameters relevant for quality assessment (QA). Based on this protocol, a round robin test was started in order to validate the genotyping results and to document the efficiency and high quality standards of the National Genotyping Platform. All laboratories of the National Genotyping platform participate in this round robin test.

The first phase is aimed to check the quality of genotyping that is performed with array-based technologies such like GeneChip™ microarrays from Affymetrix or BeadChip™ arrays from Illumina. These microarrays use a fixed set of markers that cannot be modified by the investigator. They allow the simultaneous interrogation of 100,000 – 500,000 SNPs in a single experiment from as little as 250-750 ng DNA of a sample.
Each centre using one of these high throughput chip technologies will genotype three DNA samples of a defined amount and concentration. The DNA samples are provided by the Institute for Experimental Hematology and Transfusion Medicine (IEHT). The DNA samples were anonymised and archived in the IEHT. The quality of the DNA was assessed before use. The image files of the respective quality control check (spectrum and gel electrophoresis) were attached to each DNA sample.
The genotyping results are to be sent to the center of excellence for Genetic Epidemiological Methods (GEM) in Bonn. They will be evaluated with respect to reproducibility, speed and genotyping rate. This quality assurance check will be repeated biannually. 

In a second step the quality of genotyping using flexible gene chip technology will be validated. In this phase of the round robin test 10 DNA samples will be genotyped for 10 control SNPs that were selected to meet the criterion of being also present also in high-plex panels. One of these SNPs is a chromosomal marker. These control SNPS are to be genotyped for each experiment in order to avoid a mix-up of samples.

In addition an NGFN wide control sample (CEPH-DNA) was adopted that is to be genotyped once per chips for all experiments using chip-based panels (CEPH-DNA). The genotype of this control-DNA will be archived and evaluated.

The exchange of material and data is the most important functionality of the interface between cooperating projects. As a further tool for quality assurance the use of standardized 2D-barcodes (universal identification mark) was adopted. These codes are machine readable and additionally provide for a „human readable interpretation“ (HRI) conforming to the cited ISO/IEC standards. Thereby documenting and tracing of one person’s multiple DNA-samples during the multi-layer process of genotyping and data analysis will be facilitated. For more information, download the diagram "NGFN: Identification of Probands and Material".


 Websites of institutes and centres involved in this NGFN quality management project: