Computational protocol: An unusually high frequency of SCAD deficiency caused by two pathogenic variants in the ACADS gene and its relationship to the ethnic structure in Slovakia

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Protocol publication

[…] Genomic DNA was extracted from peripheral blood leukocytes using NucleoSpin® Blood (Macherey-Nagel) according to the manufacturer’s protocol. Applying the recommendation from GeneReviews® for the molecular genetic algorithm for SCADD diagnosis, we primarily focused our study on two susceptibility variants, c.625G>A and c.511C>T, and one missense pathogenic variant, c.319C>T []. We decided to perform sequence analyses because relevant variants were found only in three biochemically defined SCADD individuals. The entire coding sequence of the ACADS gene was amplified by polymerase chain reaction (PCR) using 10 pairs of primers designed by our laboratory using the Primer3 software. Purified PCR products were sequenced using the ABI 3100® Avant Genetic Analyzer (Applied Biosystems) using a BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems). Mutation analysis was performed using SeqScape® Software v2.6. All identified sequence variants were confirmed by sequencing of both DNA strands. […]

Pipeline specifications

Software tools Primer3, SeqScape
Databases GeneReviews
Applications Sanger sequencing, qPCR
Organisms Homo sapiens
Diseases Deficiency Diseases, Genetic Diseases, Inborn