( Allelic discrim...)Detect Single Nucleotide Sequence Changes with Mx4000 Molecular Beacon,,,Allelic Discrimination Kits

Allelic discrimination kits with excellent specificity, sensitivity, and assay throughput

Xiuyuan Hu Lingyu Chen
Stratagene

Because of their unique hairpin conformation, molecular beacon probes can recognize single base differences, making them ideal for mutation detection and allelic discrimination. Based on the molecular beacon technology, we developed a series of Mx4000 molecular beacon allelic discrimination kits^,ff for the detection of single or multiple nucleotide changes in human genes. The five kits described in this article allow four single base substitutions to be detected in the human CCR2 (64I), Factor V (Leiden), SDF-1(3 A), and MTHFR (C677T) genes, as well as of a 32-base-pair deletion in the human CCR5 (D32) gene. Each kit includes two allele-specific molecular beacons in addition to genotype-specific DNA controls, PCR primers, and optimized target amplification buffer. The two molecular beacons are labeled with different fluorophores that emit fluorescent light at distinct optical wavelengths. As a result, three possible allelic combinations of two sequence variants (Allele 1/Allele 1, Allele 2/Allele 2, Allele 1/Allele 2) can be distinguished simultaneously in a DNA sample. The technique saves time and reduces the risk of contamination as the test is performed in closed tubes and requires no post-PCR manipulation of samples.

Single nucleotide sequence changes (substitution, deletion, or insertion) are the largest source of human DNA diversity, with an estimated frequency of 1 in 1,000 base pairs. Many of the sequence changes have been identified as the cause of monogenic disorders or are associated with a genetic predisposition to multifactorial diseases including cancer and cardiovascular and infectious diseases. The sequence changes also constitute the genetic basis for many
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