Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance
Abstract Background Anti-malarial drug resistance poses a threat to current global efforts towards control and elimination of malaria. Several methods are used in monitoring anti-malarial drug resistance. Molecular markers such as single nucleotide polymorphism (SNP) for example are increasingly bei...
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ftdoajarticles:oai:doaj.org/article:1cf4aa5d3c0e418e9bb5604ec0870fc2 2023-05-15T15:15:54+02:00 Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance Kamau Edwin Alemayehu Saba Feghali Karla C Tolbert LaDonna S Ogutu Bernard Ockenhouse Christian F 2012-01-01T00:00:00Z https://doi.org/10.1186/1475-2875-11-23 https://doaj.org/article/1cf4aa5d3c0e418e9bb5604ec0870fc2 EN eng BMC http://www.malariajournal.com/content/11/1/23 https://doaj.org/toc/1475-2875 doi:10.1186/1475-2875-11-23 1475-2875 https://doaj.org/article/1cf4aa5d3c0e418e9bb5604ec0870fc2 Malaria Journal, Vol 11, Iss 1, p 23 (2012) Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 article 2012 ftdoajarticles https://doi.org/10.1186/1475-2875-11-23 2022-12-31T08:18:20Z Abstract Background Anti-malarial drug resistance poses a threat to current global efforts towards control and elimination of malaria. Several methods are used in monitoring anti-malarial drug resistance. Molecular markers such as single nucleotide polymorphism (SNP) for example are increasingly being used to identify genetic mutations related to anti-malarial drug resistance. Several methods are currently being used in analysis of SNP associated with anti-malarial drug resistance and although each one of these methods has unique strengths and shortcoming, there is still need to improve and/or develop new methods that will close the gap found in the current methods. Methods TaqMan Allelic Discrimination assays for detection of SNPs associated with anti-malarial drug resistance were designed for analysis on Applied Biosystems PCR platform. These assays were designed by submitting SNP sequences associated with anti-malarial drug resistance to Applied Biosystems website. Eleven SNPs associated with resistance to anti-malarial drugs were selected and tested. The performance of each SNP assay was tested by creating plasmid DNAs carrying codons of interests and analysing them for analysis. To test the sensitivity and specificity of each SNP assay, 12 clinical samples were sequenced at codons of interest and used in the analysis. Plasmid DNAs were used to establish the Limit of Detection (LoD) for each assay. Results Data from genetic profiles of the Plasmodium falciparum laboratory strains and sequence data from 12 clinical samples was used as the reference method with which the performance of the SNP assays were compared to. The sensitivity and specificity of each SNP assay was establish at 100%. LoD for each assay was established at 2 GE, equivalent to less than 1 parasite/μL. SNP assays performed well in detecting mixed infection and analysis of clinical samples. Conclusion TaqMan Allelic Discrimination assay provides a good alternative tool in detection of SNPs associated with anti-malarial drug. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic Malaria Journal 11 1 |
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Directory of Open Access Journals: DOAJ Articles |
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English |
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Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 |
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Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 Kamau Edwin Alemayehu Saba Feghali Karla C Tolbert LaDonna S Ogutu Bernard Ockenhouse Christian F Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 Infectious and parasitic diseases RC109-216 |
description |
Abstract Background Anti-malarial drug resistance poses a threat to current global efforts towards control and elimination of malaria. Several methods are used in monitoring anti-malarial drug resistance. Molecular markers such as single nucleotide polymorphism (SNP) for example are increasingly being used to identify genetic mutations related to anti-malarial drug resistance. Several methods are currently being used in analysis of SNP associated with anti-malarial drug resistance and although each one of these methods has unique strengths and shortcoming, there is still need to improve and/or develop new methods that will close the gap found in the current methods. Methods TaqMan Allelic Discrimination assays for detection of SNPs associated with anti-malarial drug resistance were designed for analysis on Applied Biosystems PCR platform. These assays were designed by submitting SNP sequences associated with anti-malarial drug resistance to Applied Biosystems website. Eleven SNPs associated with resistance to anti-malarial drugs were selected and tested. The performance of each SNP assay was tested by creating plasmid DNAs carrying codons of interests and analysing them for analysis. To test the sensitivity and specificity of each SNP assay, 12 clinical samples were sequenced at codons of interest and used in the analysis. Plasmid DNAs were used to establish the Limit of Detection (LoD) for each assay. Results Data from genetic profiles of the Plasmodium falciparum laboratory strains and sequence data from 12 clinical samples was used as the reference method with which the performance of the SNP assays were compared to. The sensitivity and specificity of each SNP assay was establish at 100%. LoD for each assay was established at 2 GE, equivalent to less than 1 parasite/μL. SNP assays performed well in detecting mixed infection and analysis of clinical samples. Conclusion TaqMan Allelic Discrimination assay provides a good alternative tool in detection of SNPs associated with anti-malarial drug. |
format |
Article in Journal/Newspaper |
author |
Kamau Edwin Alemayehu Saba Feghali Karla C Tolbert LaDonna S Ogutu Bernard Ockenhouse Christian F |
author_facet |
Kamau Edwin Alemayehu Saba Feghali Karla C Tolbert LaDonna S Ogutu Bernard Ockenhouse Christian F |
author_sort |
Kamau Edwin |
title |
Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
title_short |
Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
title_full |
Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
title_fullStr |
Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
title_full_unstemmed |
Development of a TaqMan Allelic Discrimination Assay for detection of Single Nucleotides Polymorphisms associated with anti-malarial drug resistance |
title_sort |
development of a taqman allelic discrimination assay for detection of single nucleotides polymorphisms associated with anti-malarial drug resistance |
publisher |
BMC |
publishDate |
2012 |
url |
https://doi.org/10.1186/1475-2875-11-23 https://doaj.org/article/1cf4aa5d3c0e418e9bb5604ec0870fc2 |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
Malaria Journal, Vol 11, Iss 1, p 23 (2012) |
op_relation |
http://www.malariajournal.com/content/11/1/23 https://doaj.org/toc/1475-2875 doi:10.1186/1475-2875-11-23 1475-2875 https://doaj.org/article/1cf4aa5d3c0e418e9bb5604ec0870fc2 |
op_doi |
https://doi.org/10.1186/1475-2875-11-23 |
container_title |
Malaria Journal |
container_volume |
11 |
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1 |
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1766346230602924032 |