RT info:eu-repo/semantics/article
T1 Real-time PCR detection of PI*S and PI*Z alleles of SERPINA1 gene using SYBR green
A1 González Carracedo, Mario Andrés
A1 Ramos Díaz, Ruth
A1 Escuela Escobar, Ainhoa
A1 Díaz Usera, Ana
A1 Hernández Pérez, José María
A1 Pérez Pérez, José Antonio
A2 BioquímicaMicrobiología, Biología Celular y Genética
K1 SERPINA1 gene
K1 Alpha-1 antitrypsin deficiency
K1 PI*Z/PI*S alleles
K1 Genotyping
K1 Real-time PCR
K1 Allele-specific PCR
AB Background: Alpha-1 antitrypsin deficiency is an underdiagnosed genetic condition that predisposes to pulmonary complications and is mainly caused by rs28929474 (PI*Z allele) and rs17580 (PI*S allele) mutations in the SERPINA1 gene. Objective: Development of a homogeneous genotyping test for detection of PI*S and PI*Z alleles based on the principles of allele-specific PCR and amplicon melting analysis with a fluorescent dye. Methods: Sixty individuals, which included all possible genotypes that result from combinations of rs28929474 and rs17580 single nucleotide variants, were assayed with tailed allele-specific primers and SYBR Green dye in a real-time PCR machine. Results: A clear discrimination of mutant and wild-type variants was achieved in the genetic loci that define PI*S and PI*Z alleles. Specific amplicons showed a difference of 2.0 °C in melting temperature for non-S and S variants and of 2.9 °C for non-Z and Z variants. Conclusions: The developed genotyping method is robust, fast, and easily scalable on a standard real-time PCR platform. While it overcomes the handicaps of non-homogeneous approaches, it greatly reduces genotyping costs compared with other homogeneous approaches.
SN 0378-1119
YR 2024
FD 2024
LK http://riull.ull.es/xmlui/handle/915/41769
UL http://riull.ull.es/xmlui/handle/915/41769
LA en
NO https://portalciencia.ull.es/documentos/664b2a1acfa2e63241042711
DS Repositorio institucional de la Universidad de La Laguna
RD 18-nov-2025