Refining the results of a whole‐genome screen based on 4666 microsatellite markers for defining predisposition factors for multiple sclerosis

Abstract

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system with a complex genetic background. In order to identify loci associated with the disease, we had performed a genome screen initially using 6000 microsatellite markers in pooled DNA samples of 198 MS patients and 198 controls. Here, we report on the detailed reanalysis of this set of data. Distinctive features of microsatellites genotyped in pooled DNA causing false-positive association or masking existing association were met by improved evaluation and refined correction factors in the statistical analyses. In order to assess potential errors introduced by DNA pooling and genotyping, we resurveyed the experiment in a subset of microsatellite markers using de novo-composed DNA pools. True MS associations of markers were verified via genotyping all individual DNA samples comprised in the pools. Microsatellites share characteristically superb information content but they do not lend themselves to automation in very large scale formats. Especially after DNA pooling many artifacts of individual marker systems require special attention and treatment. Therefore, in the near future comprehensive whole-genome screens may rather be performed by typing single nucleotide polymorphisms on chip-based platforms.

SOX9 Duplication Linked to Intersex in Deer

“…Fragment analyses were performed as described previously [24] on a Beckman-Coulter CEQ 8000 capillary sequencer following standard protocols and using the included software. The described primer sequences from Pajares et al (2007) were optimized under our own conditions.…”

An extended association screen in multiple sclerosis using 202 microsatellite markers targeting apoptosis-related genes does not reveal new predisposing factors

“…For the NFκB2 , NGFβ and NOS1 genes, the comparisons of allele frequencies from pooled and individually-typed DNA samples (Fig. 1) show an important and typical artifact [9]. DNA polymerases tend to preferentially amplify short alleles in favour of longer alleles (length-dependent amplification).…”

“…The employment of pooled DNA samples in microsatellite genotyping introduces errors [9], unless pooling is performed absolutely accurately. Concentration of DNA from each individual was quantified in triplicate using spectrophotometric measurement and then diluted to a final 50 ng/μl.…”

Functionally relevant variations of the interleukin‐10 gene associated with antineutrophil cytoplasmic antibody–negative Churg‐Strauss syndrome, but not with Wegener's granulomatosis

“…Five hundred seven healthy blood donors from Germany (274 male, 233 female) were used as controls. This control group was previously described in an earlier genetic study (43), and ancestry of the control subjects was evaluated in the same manner as for the patient group. This control group has already been used in numerous genetic association studies of (auto)immune disorders, such as multiple sclerosis (40) and WG (13, 14, 40, 41).…”