ArticleNucleic acids research2019
Engineering of a thermostable viral polymerase using metagenome-derived diversity for highly sensitive and specific RT-PCR.
Article in Nucleic acids research, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 12 papers, 1 of them a synthesis that pooled it.
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Who cites it
12 citing papers in PubMed, 1 synthesis or guideline pooled it, 28 citations in OpenAlex.
- Bacteriophages of Thermophilic 'International journal of molecular sciences · 2024Pooled it
- Engineering of novel DNA polymerase variants for single enzyme quantitative multiplex reverse transcription-PCR.Scientific reports · 2025Article
- Comparison of Commercially Available Thermostable DNA Polymerases with Reverse Transcriptase Activity in Coupled Reverse Transcription Polymerase Chain Reaction Assays.Methods and protocols · 2025Article
- A Role of Multi-Omics Technologies in Sheep and Goat Meats: Progress and Way Ahead.Foods (Basel, Switzerland) · 2023Review
- Multi-Omics Approaches to Improve Meat Quality and Taste Characteristics.Food science of animal resources · 2023Review
- Functional biology and biotechnology of thermophilic viruses.Essays in biochemistry · 2023Review
- Potential Applications of Thermophilic Bacteriophages in One Health.International journal of molecular sciences · 2023Review
- RT-qPCR Detection of SARS-CoV-2: No Need for a Dedicated Reverse Transcription Step.International journal of molecular sciences · 2022Article
- Rational-Design Engineering to Improve Enzyme Thermostability.Methods in molecular biology (Clifton, N.J.) · 2022Article
- Review
- Diversity and Distribution of a Novel Genus of HyperthermophilicFrontiers in microbiology · 2020Article
- A Review on Viral Metagenomics in Extreme Environments.Frontiers in microbiology · 2019Review
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
6 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Reverse transcription is an essential initial step in the analysis of RNA for most PCR-based amplification and detection methods. Despite advancements in these technologies, efficient conversion of RNAs that form stable secondary structures and double-stranded RNA targets remains challenging as retroviral-derived reverse transcriptases are often not sufficiently thermostable to catalyze synthesis at temperatures high enough to completely relax these structures. Here we describe the engineering and improvement of a thermostable viral family A polymerase with inherent reverse transcriptase activity for use in RT-PCR. Using the 3173 PyroPhage polymerase, previously identified from hot spring metagenomic sampling, and additional thermostable orthologs as a source of natural diversity, we used gene shuffling for library generation and screened for novel variants that retain high thermostability and display elevated reverse transcriptase activity. We then created a fusion enzyme between a high-performing variant polymerase and the 5'→3' nuclease domain of Taq DNA polymerase that provided compatibility with probe-based detection chemistries and enabled highly sensitive detection of structured RNA targets. This technology enables a flexible single-enzyme RT-PCR system that has several advantages compared with standard heat-labile reverse transcription methods.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.