Real-time PCR detection of PI*S and PI*Z alleles of SERPINA1 gene using SYBR green.
Gene
; 921: 148540, 2024 Aug 30.
Article
em En
| MEDLINE
| ID: mdl-38723785
ABSTRACT
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.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Compostos Orgânicos
/
Quinolinas
/
Alfa 1-Antitripsina
/
Diaminas
/
Alelos
/
Benzotiazóis
/
Reação em Cadeia da Polimerase em Tempo Real
Limite:
Humans
Idioma:
En
Revista:
Gene
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Espanha