Genotyping Microarray for the Detection of More Than 200 CFTR Mutations in Ethnically Diverse Populations

Abstract

Cystic fibrosis (CF), which is due to mutations in the cystic fibrosis transmembrane conductance regulator gene, is a common life-shortening disease. Although CF occurs with the highest incidence in Caucasians, it also occurs in other ethnicities with variable frequency. Recent national guidelines suggest that all couples contemplating pregnancy should be informed of molecular screening for CF carrier status for purposes of genetic counseling. Commercially available CF carrier screening panels offer a limited panel of mutations, however, making them insufficiently sensitive for certain groups within an ethnically diverse population. This discrepancy is even more pronounced when such carrier screening panels are used for diagnostic purposes. By means of arrayed primer extension technology, we have designed a genotyping microarray with 204 probe sites for CF transmembrane conductance regulator gene mutation detection. The arrayed primer extension array, based on a platform technology for disease detection with multiple applications, is a robust, costeffective, and easily modifiable assay suitable for CF carrier screening and disease detection. (J Mol Diagn 2005, 7:375-387)

Assessing human germ‐cell mutagenesis in the Postgenome Era: A celebration of the legacy of William Lawson (Bill) Russell

“…Highthroughput molecular genetics has changed the face of medical research dramatically, but it has perhaps had less of an impact on clinical medicine than one might expect [Tischfield et al, 2005]. For example, it is technically feasible to use gene-chip methodology to screen for any of several hundred possible mutations that cause cystic fibrosis [Schrijver et al, 2005]. However, the logistics of gene-chip development are difficult.…”

Two-dye based arrayed primer extension for simultaneous multigene detection in lipid metabolism

“…The APEX microarray has been developed over a decade [26] and mostly applied to diagnose genetic disease [27][28][29][30]. APEX is based on a two-step reaction; the hybridization between probe and target, and a single base extension by DNA polymerase at the polymorphic site with four unique dye-labeled dideoxy nucleotides [23,[27][28][29][30].…”

“…APEX is based on a two-step reaction; the hybridization between probe and target, and a single base extension by DNA polymerase at the polymorphic site with four unique dye-labeled dideoxy nucleotides [23,[27][28][29][30]. The APEX assay is robust, cost-effective, and easily modified, which provides a higher power of identification of mutations (missense, deletion, insertion) or polymorphisms in heterozygous and homozygous patients when compared with other common microarray technologies [23].…”

Comprehensive Arrayed Primer Extension Array for the Detection of 59 Sequence Variants in 15 Conditions Prevalent Among the (Ashkenazi) Jewish Population

“…34 For each mutation under interrogation, two forward and two reverse strand oligonucleotides were spotted, for a total of four data points per possible sequence variant. To reduce the background fluorescence and to avoid rehybridization of unbound oligonucleotides to the APEX slide, the slides were washed with 95°C distilled water and 100 mmol/L NaOH before the APEX reactions.…”