Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene
Background: Expansion of GAA trinucleotide repeats is the molecular basis of Friedreich’s ataxia (FRDA). Precise detection of the GAA expansion repeat in frataxin gene has always been a challenge. Different molecular methods have been suggested for detection of GAA expansion, including; short-PCR, l...
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| Format: | Article |
| Language: | English |
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Tehran University of Medical Sciences
2017-02-01
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| Series: | Iranian Journal of Public Health |
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| Online Access: | https://ijph.tums.ac.ir/index.php/ijph/article/view/9052 |
| _version_ | 1811190150548422656 |
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| author | Mona ENTEZAM Akbar AMIRFIROOZI Mansoureh TOGHA Mohammad KERAMATIPOUR |
| author_facet | Mona ENTEZAM Akbar AMIRFIROOZI Mansoureh TOGHA Mohammad KERAMATIPOUR |
| author_sort | Mona ENTEZAM |
| collection | DOAJ |
| description | Background: Expansion of GAA trinucleotide repeats is the molecular basis of Friedreich’s ataxia (FRDA). Precise detection of the GAA expansion repeat in frataxin gene has always been a challenge. Different molecular methods have been suggested for detection of GAA expansion, including; short-PCR, long-PCR, Triplet repeat primed-PCR (TP-PCR) and southern blotting. The aim of study was to evaluate two PCR-based methods, TP-PCR and long-PCR, and to explore the use of TP-PCR accompanying with long-PCR for accurate genotyping of FRDA patients.
Methods: Blood samples were collected from six Iranian patients suspected to FRDA, who referred to the Department of Medical Genetics at Tehran University of Medical Sciences during the year 2014. For one of these patients’ four asymptomatic members of the family were also recruited for the analysis. DNA extraction was performed by two different methods. TP-PCR and long-PCR were carried out in all samples. The type of this study is assessment / investigation of methods.
Results: Using a combination of the above methods, the genotypes of all samples were confirmed as five homozygous mutants (expanded GAA repeats), two heterozygous and three homozygous normal (normal repeat size). The results obtained by TP-PCR are consistent with long-PCR results.
Conclusion: The presence or absence of expanded alleles can be identified correctly by TP-PCR. Performing long-PCR and Fluorescent-long-PCR enables accurate genotyping in all samples. This approach is highly reliable. It could be successfully used for detection of GAA expansion repeats. |
| first_indexed | 2024-04-11T14:45:30Z |
| format | Article |
| id | doaj.art-e008cec43cfa4f2db5204873739f2b3a |
| institution | Directory Open Access Journal |
| issn | 2251-6085 2251-6093 |
| language | English |
| last_indexed | 2024-04-11T14:45:30Z |
| publishDate | 2017-02-01 |
| publisher | Tehran University of Medical Sciences |
| record_format | Article |
| series | Iranian Journal of Public Health |
| spelling | doaj.art-e008cec43cfa4f2db5204873739f2b3a2022-12-22T04:17:40ZengTehran University of Medical SciencesIranian Journal of Public Health2251-60852251-60932017-02-01462Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin GeneMona ENTEZAM0Akbar AMIRFIROOZI1Mansoureh TOGHA2Mohammad KERAMATIPOUR3Dept. of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, IranDept. of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, IranIranian Center of Neurological Research, Neuroscience Institute, Tehran University of Medical Sciences, Tehran, Iran AND Dept. of Neurology, Sina Hospital, Tehran University of Medical Sciences, Tehran, IranDept. of Medical Genetics, School of Medicine, Tehran University of Medical Sciences, Tehran, IranBackground: Expansion of GAA trinucleotide repeats is the molecular basis of Friedreich’s ataxia (FRDA). Precise detection of the GAA expansion repeat in frataxin gene has always been a challenge. Different molecular methods have been suggested for detection of GAA expansion, including; short-PCR, long-PCR, Triplet repeat primed-PCR (TP-PCR) and southern blotting. The aim of study was to evaluate two PCR-based methods, TP-PCR and long-PCR, and to explore the use of TP-PCR accompanying with long-PCR for accurate genotyping of FRDA patients. Methods: Blood samples were collected from six Iranian patients suspected to FRDA, who referred to the Department of Medical Genetics at Tehran University of Medical Sciences during the year 2014. For one of these patients’ four asymptomatic members of the family were also recruited for the analysis. DNA extraction was performed by two different methods. TP-PCR and long-PCR were carried out in all samples. The type of this study is assessment / investigation of methods. Results: Using a combination of the above methods, the genotypes of all samples were confirmed as five homozygous mutants (expanded GAA repeats), two heterozygous and three homozygous normal (normal repeat size). The results obtained by TP-PCR are consistent with long-PCR results. Conclusion: The presence or absence of expanded alleles can be identified correctly by TP-PCR. Performing long-PCR and Fluorescent-long-PCR enables accurate genotyping in all samples. This approach is highly reliable. It could be successfully used for detection of GAA expansion repeats.https://ijph.tums.ac.ir/index.php/ijph/article/view/9052Triplet repeat primed-PCRLong-PCRGAA trinucleotide repeatFriedreich’s ataxiaFrataxin (FXN) gene |
| spellingShingle | Mona ENTEZAM Akbar AMIRFIROOZI Mansoureh TOGHA Mohammad KERAMATIPOUR Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene Iranian Journal of Public Health Triplet repeat primed-PCR Long-PCR GAA trinucleotide repeat Friedreich’s ataxia Frataxin (FXN) gene |
| title | Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene |
| title_full | Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene |
| title_fullStr | Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene |
| title_full_unstemmed | Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene |
| title_short | Comparison of Two Different PCR-based Methods for Detection of GAA Expansions in Frataxin Gene |
| title_sort | comparison of two different pcr based methods for detection of gaa expansions in frataxin gene |
| topic | Triplet repeat primed-PCR Long-PCR GAA trinucleotide repeat Friedreich’s ataxia Frataxin (FXN) gene |
| url | https://ijph.tums.ac.ir/index.php/ijph/article/view/9052 |
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