Real time PCR detection of the PI″Z and PI″S mutations associated with alpha-1 antitrypsin deficiency

Claudine L. Bartels, Angela L. Marchetti, W Edward Jr. Highsmith, Gregory J. Tsongalis

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Alpha-1 antitrypsin (A1AT or AAT) is a serine protease inhibitor (PI) which, when present at low levels, can cause chronic obstructive pulmonary disease (COPD) and liver disease in both children and adults. Several mutations within the SERPINA1 gene have been found to cause this deficiency. The most common variants are PI Z and PI S, each caused by a single nucleotide polymorphism (SNP). We describe a real time polymerase chain reaction (PCR) assay for the rapid genotyping of these polymorphisms. DNA was extracted from fourteen EDTA-anticoagulated whole blood samples using the Qiagen EZ1 blood extraction kit. SNP genotyping was performed using primer/probe sets purchased from Applied Biosystems. These were evaluated for performance and assay conditions on the Applied Biosystems 7500 FAST System. The genotypes of these samples were compared with their phenotype results from isoelectric focusing assays, which were performed by an independent reference laboratory. In addition, twenty samples that were previously genotyped at another laboratory were obtained for accuracy studies. Thirty-four samples were tested; five genotypes were represented and the assay was able to discriminate these successfully. Only one genotype could not be correlated with its phenotype result, as the phenotype was reported as an "unidentified allele". All other genotyping results were concordant with previously determined genotypes and phenotypes. We describe a rapid real time PCR assay that is suitable for clinical use in genotyping AAT alleles and which can be used as the initial step in A1AT testing algorithms.

Original languageEnglish (US)
Pages (from-to)406-411
Number of pages6
JournalAmerican Journal of Translational Research
Volume1
Issue number4
StatePublished - 2009

Fingerprint

alpha 1-Antitrypsin Deficiency
alpha 1-Antitrypsin
Polymerase chain reaction
Real-Time Polymerase Chain Reaction
Assays
Genotype
Phenotype
Mutation
Polymorphism
Protease Inhibitors
Single Nucleotide Polymorphism
Alleles
Blood
Nucleotides
Serine Proteinase Inhibitors
Isoelectric Focusing
Pulmonary diseases
Edetic Acid
Chronic Obstructive Pulmonary Disease
Liver Diseases

Keywords

  • Alpha-1 antitrypsin
  • Chronic obstructive pulmonary disease (COPD)
  • Genotyping
  • Molecular diagnostics
  • Real-time PCR
  • Serine protease inhibitor
  • Single nucleotide polymorphism (SNP)

ASJC Scopus subject areas

  • Medicine(all)
  • Cancer Research
  • Clinical Biochemistry
  • Molecular Medicine

Cite this

Bartels, C. L., Marchetti, A. L., Highsmith, W. E. J., & Tsongalis, G. J. (2009). Real time PCR detection of the PI″Z and PI″S mutations associated with alpha-1 antitrypsin deficiency. American Journal of Translational Research, 1(4), 406-411.

Real time PCR detection of the PI″Z and PI″S mutations associated with alpha-1 antitrypsin deficiency. / Bartels, Claudine L.; Marchetti, Angela L.; Highsmith, W Edward Jr.; Tsongalis, Gregory J.

In: American Journal of Translational Research, Vol. 1, No. 4, 2009, p. 406-411.

Research output: Contribution to journalArticle

Bartels, CL, Marchetti, AL, Highsmith, WEJ & Tsongalis, GJ 2009, 'Real time PCR detection of the PI″Z and PI″S mutations associated with alpha-1 antitrypsin deficiency', American Journal of Translational Research, vol. 1, no. 4, pp. 406-411.
Bartels, Claudine L. ; Marchetti, Angela L. ; Highsmith, W Edward Jr. ; Tsongalis, Gregory J. / Real time PCR detection of the PI″Z and PI″S mutations associated with alpha-1 antitrypsin deficiency. In: American Journal of Translational Research. 2009 ; Vol. 1, No. 4. pp. 406-411.
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