Mahmoud Zhra, Aljohara Al Saud, Maha Alzayer, Liliane Okdah, Hani Tamim, Hana M. A. Fakhoury, Ahmad Aljada
{"title":"Cost-effective in-house COVID-19 reverse transcription-polymerase chain reaction testing with yeast-derived Taq polymerase","authors":"Mahmoud Zhra, Aljohara Al Saud, Maha Alzayer, Liliane Okdah, Hani Tamim, Hana M. A. Fakhoury, Ahmad Aljada","doi":"10.4103/atm.atm_180_23","DOIUrl":null,"url":null,"abstract":"\n \n Despite the decline of the COVID-19 pandemic, there continues to be a persistent requirement for reliable testing methods that can be adapted to future outbreaks and areas with limited resources. While the standard approach of using reverse transcription-polymerase chain reaction (RT-PCR) with Taq polymerase is effective, it faces challenges such as limited access to high-quality enzymes and the presence of bacterial DNA contamination in commercial kits, which can impact the accuracy of test results.\n \n \n \n This study investigates the production of recombinant Taq polymerase in yeast cells and assesses its crude lysate in a multiplex RT-PCR assay for detecting the SARS-CoV-2 RNA-dependent RNA polymerase (RdRP) and N genes, with human Ribonuclease P serving as an internal control.\n \n \n \n The unpurified yeast Taq polymerase demonstrates sensitivity comparable to commercially purified bacterial Taq polymerase and unpurified bacterial counterparts in detecting the RdRP and N genes. It exhibits the highest specificity, with 100% accuracy, for the N gene. The specificity for the RdRP gene closely aligns with that of commercially purified bacterial Taq polymerase and unpurified bacterial Taq polymerase.\n \n \n \n The use of unpurified recombinant yeast Taq polymerase shows promise as a cost-effective approach for conducting in-house COVID-19 RT-PCR testing. By eliminating the need for chromatography purification steps, the production of RT-PCR kits can be streamlined, potentially improving accessibility and scalability, especially in resource-limited settings and future pandemics.\n","PeriodicalId":50760,"journal":{"name":"Annals of Thoracic Medicine","volume":null,"pages":null},"PeriodicalIF":2.1000,"publicationDate":"2024-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Annals of Thoracic Medicine","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.4103/atm.atm_180_23","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"CARDIAC & CARDIOVASCULAR SYSTEMS","Score":null,"Total":0}
引用次数: 0
Abstract
Despite the decline of the COVID-19 pandemic, there continues to be a persistent requirement for reliable testing methods that can be adapted to future outbreaks and areas with limited resources. While the standard approach of using reverse transcription-polymerase chain reaction (RT-PCR) with Taq polymerase is effective, it faces challenges such as limited access to high-quality enzymes and the presence of bacterial DNA contamination in commercial kits, which can impact the accuracy of test results.
This study investigates the production of recombinant Taq polymerase in yeast cells and assesses its crude lysate in a multiplex RT-PCR assay for detecting the SARS-CoV-2 RNA-dependent RNA polymerase (RdRP) and N genes, with human Ribonuclease P serving as an internal control.
The unpurified yeast Taq polymerase demonstrates sensitivity comparable to commercially purified bacterial Taq polymerase and unpurified bacterial counterparts in detecting the RdRP and N genes. It exhibits the highest specificity, with 100% accuracy, for the N gene. The specificity for the RdRP gene closely aligns with that of commercially purified bacterial Taq polymerase and unpurified bacterial Taq polymerase.
The use of unpurified recombinant yeast Taq polymerase shows promise as a cost-effective approach for conducting in-house COVID-19 RT-PCR testing. By eliminating the need for chromatography purification steps, the production of RT-PCR kits can be streamlined, potentially improving accessibility and scalability, especially in resource-limited settings and future pandemics.
期刊介绍:
The journal will cover studies related to multidisciplinary specialties of chest medicine, such as adult and pediatrics pulmonology, thoracic surgery, critical care medicine, respiratory care, transplantation, sleep medicine, related basic medical sciences, and more. The journal also features basic science, special reports, case reports, board review , and more. Editorials and communications to the editor that explore controversial issues and encourage further discussion by physicians dealing with chest medicine.