ArticleExtremophiles : life under extreme conditions2015
Synergistic template-free synthesis of dsDNA by Thermococcus nautili primase PolpTN2, DNA polymerase PolB, and pTN2 helicase.
Article in Extremophiles : life under extreme conditions, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 5 papers.
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5 citing papers in PubMed, 8 citations in OpenAlex.
- A primer-independent DNA polymerase-based method for competent whole-genome amplification of intermediate to high GC sequences.NAR genomics and bioinformatics · 2023Article
- The primordial tRNA acceptor stem code from theoretical minimal RNA ring clusters.BMC genetics · 2020Article
- Systematic Nucleotide Exchange Analysis of ESTs From the Human Cancer Genome Project Report: Origins of 347 Unknown ESTs Indicate Putative Transcription of Non-Coding Genomic Regions.Frontiers in genetics · 2020Article
- Viruses and Evolution - Viruses First? A Personal Perspective.Frontiers in microbiology · 2019Article
- Stem-Loop RNA Hairpins in Giant Viruses: Invading rRNA-Like Repeats and a Template Free RNA.Frontiers in microbiology · 2018Article
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Authors and funding
4 authors at 1 institution in 1 country.
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Abstract
A combination of three enzymes from the hyperthermophilic archaeon Thermococcus nautili, DNA primase PolpTN2, DNA polymerase PolB, and pTN2 DNA helicase, was found to synthesize up to 300-400 ng/µl dsDNA from deoxynucleotide triphosphates in less than 30 min in the absence of added template DNA and oligonucleotide primer. The reaction did not occur below 64 °C. No synthesis was observed if PolpTN2 or PolB were left out; helicase was not essential but accelerated the reaction. The DNA synthesized consisted of highly reiterated palindromic sequences reaching up to more that 10 kb. Sequence analysis of three independent reaction products synthesized at different temperatures showed that the palindromes shared a common pentanucleotide core, suggesting that random nucleic acid fragments were not responsible for priming the reaction. When enzymes were added sequentially, preincubation with primase plus helicase followed by PolB led to a shorter delay before the onset of the reaction as compared to preincubation with PolB plus helicase followed by primase. This suggests that the primase generates seeds that are subsequently amplified and elongated in synergy with PolB by a mechanism involving hairpin formation and slippage synthesis.
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