The influence of quality of primers on the formation of primer dimers in PCR

Гарафутдинов, Р.Р.; Galimova, A. A.; Сахабутдинова, А. Р.

doi:10.1080/15257770.2020.1803354

Cited by 11 publications

(16 citation statements)

References 37 publications

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“…When using DNA #1, #2, #3, #4, #5, and #6 at low concentrations of less than 10 5 molecules, nonspecific amplification such as primer dimers was observed, and we did not obtain linear calibration curves. Even primers designed using mathematical models sometimes form nonspecific products in negative controls and require PCR optimization. , …”

Section: Resultsmentioning

confidence: 99%

“…Even primers designed using mathematical models sometimes form nonspecific products in negative controls and require PCR optimization. 37,38 As an initial trial, the nanopore filter experiment was performed using six different types of ssDNA, DNA #1−6, under conventional conditions for nanopore sensing. Clear blocking signals of ssDNA (160 nM) translocation events were observed for the buffer condition of 1 M KCl, 10 mM MOPS, pH 7.0.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

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Nanopore Filter: A Method for Counting and Extracting Single DNA Molecules Using a Biological Nanopore

et al. 2023

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This paper describes a method for the real-time counting and extraction of DNA molecules at the single-molecule level by nanopore technology. As a powerful tool for electrochemical single-molecule detection, nanopore technology eliminates the need for labeling or partitioning sample solutions at the femtoliter level. Here, we attempt to develop a DNA filtering system utilizing an α-hemolysin (αHL) nanopore. This system comprises two droplets, one filling with and one emptying DNA molecules, separated by a planar lipid bilayer containing αHL nanopores. The translocation of DNA through the nanopores is observed by measuring the channel current, and the number of translocated molecules can also be verified by quantitative polymerase chain reaction (qPCR). However, we found that the issue of contamination seems to be an almost insolvable problem in single-molecule counting. To tackle this problem, we tried to optimize the experimental environment, reduce the volume of solution containing the target molecule, and use the PCR clamp method. Although further efforts are still needed to achieve a singlemolecule filter with electrical counting, our proposed method shows a linear relationship between the electrical counting and qPCR estimation of the number of DNA molecules.

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Section: Resultsmentioning

confidence: 99%

Section: ■ Results and Discussionmentioning

confidence: 99%

Nanopore Filter: A Method for Counting and Extracting Single DNA Molecules Using a Biological Nanopore

et al. 2023

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show abstract

“…Surprisingly, a 10-fold reduction of the enzyme amount resulted in a significant decrease of the threshold cycle (Ct) value (∼6 cycles) for RNA-positive samples. A 2-fold decrease of the primers and dNTP concentration decreased Ct values as well, probably due to the suppression of nonspecific amplification caused by the competition between specific and nonspecific DNA synthesis [ 35 ].…”

Section: Resultsmentioning

confidence: 99%

Reverse transcriptase-free detection of viral RNA using Hemo Klentaq DNA polymerase

Сахабутдинова

Gazizov

Чемерис

et al. 2022

Analytical Biochemistry

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“…Although some subgroup analyses were conducted to identify the sources of heterogeneity, many unmeasured moderators have impacts on the results. For example, the primer sets used in the included studies varied, and the sensitivity of different primer sets was unclear [ 74 ]. Finally, similar with other meta-analyses on tick-borne infections [ 11 , 75 , 76 ], publication bias exists in the present study, which may distort the estimates of prevalence, so results should be interpreted with caution.…”

Section: Discussionmentioning

confidence: 99%

Nucleic Acid Prevalence of Zoonotic Babesia in Humans, Animals and Questing Ticks, a Systematic Review and Meta-Analysis

Yao

Tian

et al. 2023

TropicalMed

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Background: Zoonotic Babesia infections are an emerging public health threat globally. The geographical distribution, animal reservoirs and tick vectors vary greatly across Babesia species, and estimations of prevalence reported in works within the literature are also quite different. Better prevalence estimates and identification of moderators are needed to understand the global transmission risk of different zoonotic Babesia species, and to provide crucial background information for the diagnosis, treatment and control of zoonotic babesiosis. Methods: We conducted a systematic review and meta-analysis to determine the global nucleic acid prevalence of different zoonotic Babesia species in humans, animals and ticks. Relevant publications were obtained from several electronic databases and grey literature up to December 2021. Articles were included if they were published in English or Chinese and reported the nucleic acid prevalence of zoonotic Babesia species in humans, animals or ticks. The pooled estimates of prevalence were determined using a random effect model. Heterogeneity was investigated using subgroup analyses and random effect meta-regression models. Results: Of 3205 unique studies, 28 were included by the systematic review of zoonotic Babesia for humans, 79 for animals and 104 for ticks. The results showed overall pooled estimates of nucleic acid prevalence for the following: B. microti—1.93% (0.32–4.69%) in humans; B. microti—7.80% (5.25–10.77%), B. divergens—2.12% (0.73–4.08%) and B. venatorum—1.42% (0.30–3.16%) in animals; and B. microti—2.30% (1.59–3.13%), B. divergens—0.16% (0.05–0.32%), and B. venatorum—0.39% (0.26–0.54%) in questing ticks. The type of population, animal reservoir or tick vector, detecting method and continent were moderators possibly associated with heterogeneity, yet the remaining heterogeneity that was not explained was still substantial (all QE p values < 0.05). Conclusions: B. microti is the most prevalent and widely distributed zoonotic Babesia species globally. The wide range of suitable animal reservoirs and potential transmission vectors and high prevalence in animals and ticks may contribute to the worldwide distribution of B. microti. Other zoonotic Babesia species were relatively less prevalent and were reported in quite limited areas.

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The influence of quality of primers on the formation of primer dimers in PCR

Cited by 11 publications

References 37 publications

Nanopore Filter: A Method for Counting and Extracting Single DNA Molecules Using a Biological Nanopore

Nanopore Filter: A Method for Counting and Extracting Single DNA Molecules Using a Biological Nanopore

Reverse transcriptase-free detection of viral RNA using Hemo Klentaq DNA polymerase

Nucleic Acid Prevalence of Zoonotic Babesia in Humans, Animals and Questing Ticks, a Systematic Review and Meta-Analysis

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