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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 559 records · Page 31
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Structure- and Interaction-Based Design of Anti-SARS-CoV-2 Aptamers

Aptamer selection against novel infections is a complicated and time-consuming approach. Synergy can be achieved by using computational methods together with experimental procedures. In this study, we aim to develop a reliable methodology for a rational aptamer in silico et vitro design. The new approach combines multiple steps: (1) Molecular design, based on screening in a DNA aptamer library and directed mutagenesis to fit the protein tertiary structure; (2) 3D molecular modeling of the target; (3) Molecular docking of an aptamer with the protein; (4) Molecular dynamics (MD) simulations of the complexes; (5) Quantum-mechanical (QM) evaluation of the interactions between aptamer and target with further analysis; (6) Experimental verification at each cycle for structure and binding affinity by using small-angle X-ray scattering, cytometry, and fluorescence polarization. By using a new iterative design procedure, structure- and interaction-based drug design (SIBDD), a highly specific aptamer to the receptor-binding domain of the SARS-CoV-2 spike protein, was developed and validated. The SIBDD approach enhances speed of the high-affinity aptamers development from scratch, using a target protein structure. The method could be used to improve existing aptamers for stronger binding. This approach brings to an advanced level the development of novel affinity probes, functional nucleic acids. It offers a blueprint for the straightforward design of targeting molecules for new pathogen agents and emerging variants.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

CHARMM-GUI Drude prepper for molecular dynamics simulation using the classical Drude polarizable force field

Explicit treatment of electronic polarizability in empirical force fields (FFs) represents an extension over a traditional additive or pairwise FF and provides a more realistic model of the variations in electronic structure in condensed phase, macromolecular simulations. To facilitate utilization of the polarizable FF based on the classical Drude oscillator model, Drude Prepper has been developed in CHARMM-GUI. Drude Prepper ingests additive CHARMM protein structures file (PSF) and pre-equilibrated coordinates in CHARMM, PDB, or NAMD format, from which the molecular components of the system are identified. These include all residues and patches connecting those residues along with water, ions, and other solute molecules. This information is then used to construct the Drude FF-based PSF using molecular generation capabilities in CHARMM, followed by minimization and equilibration. In addition, inputs are generated for molecular dynamics (MD) simulations using CHARMM, GROMACS, NAMD, and OpenMM. Validation of the Drude Prepper protocol and inputs is performed through conversion and MD simulations of various heterogeneous systems that include proteins, nucleic acids, lipids, polysaccharides, and atomic ions using the aforementioned simulation packages. Stable simulations are obtained in all studied systems, including 5 μs simulation of ubiquitin, verifying the integrity of the generated Drude PSFs. Additionally, the ability of the Drude FF to model variations in electronic structure is shown through dipole moment analysis in selected systems. Finally, the capabilities and availability of Drude Prepper in CHARMM-GUI is anticipated to greatly facilitate the application of the Drude FF to a range of condensed phase, macromolecular systems.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2019

Metabolite composition of sinking particles differs from surface suspended particles across a latitudinal transect in the South Atlantic

Abstract Marine sinking particles transport carbon from the surface and bury it in deep‐sea sediments, where it can be sequestered on geologic time scales. The combination of the surface ocean food web that produces these particles and the particle‐associated microbial community that degrades them creates a complex set of variables that control organic matter cycling. We use targeted metabolomics to characterize a suite of small biomolecules, or metabolites, in sinking particles and compare their metabolite composition to that of the suspended particles in the euphotic zone from which they are likely derived. These samples were collected in the South Atlantic subtropical gyre, as well as in the equatorial Atlantic region and the Amazon River plume. The composition of targeted metabolites in the sinking particles was relatively similar throughout the transect, despite the distinct oceanic regions in which they were generated. Metabolites possibly derived from the degradation of nucleic acids and lipids, such as xanthine and glycine betaine, were an increased mole fraction of the targeted metabolites in the sinking particles relative to surface suspended particles, while algal‐derived metabolites like the osmolyte dimethylsulfoniopropionate were a smaller fraction of the observed metabolites on the sinking particles. These compositional changes are shaped both by the removal of metabolites associated with detritus delivered from the surface ocean and by production of metabolites by the sinking particle‐associated microbial communities. Furthermore, they provide a basis for examining the types and quantities of metabolites that may be delivered to the deep sea by sinking particles.

Johnson, Winifred M.↗
DOE OSTI2025

Visualizing and analyzing 3D biomolecular structures using Mol* at RCSB.org: Influenza A H5N1 virus proteome case study

The easiest and often most useful way to work with experimentally determined or computationally predicted structures of biomolecules is by viewing their three-dimensional (3D) shapes using a molecular visualization tool. Mol* was collaboratively developed by RCSB Protein Data Bank (RCSB PDB, RCSB.org) and Protein Data Bank in Europe (PDBe, PDBe.org) as an open-source, web-based, 3D visualization software suite for examination and analyses of biostructures. It is capable of displaying atomic coordinates and related experimental data of biomolecular structures together with a variety of annotations, facilitating basic and applied research, training, education, and information dissemination. Across RCSB.org, the RCSB PDB research-focused web portal, Mol* has been implemented to support single-mouse-click atomic-level visualization of biomolecules (e.g., proteins, nucleic acids, carbohydrates) with bound cofactors, small-molecule ligands, ions, water molecules, or other macromolecules. RCSB.org Mol* can seamlessly display 3D structures from various sources, allowing structure interrogation, superimposition, and comparison. Using influenza A H5N1 virus as a topical case study of an important pathogen, we exemplify how Mol* has been embedded within various RCSB.org tools—allowing users to view polymer sequence and structure-based annotations integrated from trusted bioinformatics data resources, assess patterns and trends in groups of structures, and view structures of any size and compositional complexity. In addition to being linked to every experimentally determined biostructure and Computed Structure Model made available at RCSB.org, Standalone Mol* is freely available for visualizing any atomic-level or multi-scale biostructure at rcsb.org/3d-view.

3D biostructure↗
DOE OSTI2023

Synthesis of Pyrimidine Modified Seleno–DNA as a Novel Approach to Antisense Candidate

Chemically modified antisense oligonucleotides (ASO) currently in pre–clinical and clinical experiments mainly focus on the 2’–position derivatizations to enhance stability and targeting affinity. Considering the possible incompatibility of 2’–modifications with RNase H stimulation and activity, we have hypothesized that the atom specific modifications on nucleobases can retain the complex structure and RNase H activity, while enhancing ASO's binding affinity, specificity, and stability against nucleases. Herein we report a novel strategy to explore our hypothesis by synthesizing the deoxynucleoside phosphoramidite building block with the seleno–modification at 5–position of thymidine, as well as its Se–oligonucleotides. Via X–ray crystal structural study, we found that the Se–modification was located in the major groove of nucleic acid duplex and didn't cause the thermal and structural perturbations. Surprisingly, our nucleobase–modified Se–DNAs were exceptionally resistant to nuclease digestion, while compatible with RNase H activity. In conclusion, this affords a novel avenue for potential anti–sense modification in the form of Se–antisense oligonucleotides (Se–ASO).

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2020

Direct SERDS sensing of molecular biomarkers in plants under field conditions

Molecular biomarkers such as microRNAs (miRNAs) play important roles in regulating various developmental processes in plants. Understanding these pathways will help bioengineer designing organisms for efficient biomass accumulation. Current methods for RNA analysis require sample extraction and multi-step sample analysis, hindering work in field studies. Recent work in the incorporation of nanomaterials for plant bioengineering research is leading the way of an agri-tech revolution. As an example, surface-enhanced Raman scattering (SERS)–based sensors can be used to monitor RNA in vivo. However, the use of SERS in the field has been limited due to issues with observing Raman signal over complex background. To this end, shifted-excitation Raman difference spectroscopy (SERDS) offers an effective solution to extract the SERS signal from high background based on a physical approach. In this manuscript, we report the first application of SERDS on SERS sensors. We investigated this technique on SERS sensor developed for the detection of a microRNA biomarker, miR858. We tested the technique on in vitro samples and validated the technique by detecting the presence of exogenous miR858 in plants directly under ambient light in a growth chamber. The possibility of moving the detection of nucleic acid targets outside the constraints of laboratory setting enables numerous important bioengineering applications. Finally, such applications can revolutionize biofuel production and agri-tech through the use of nanotechnology-based monitoring of plant growth, plant health, and exposure to pollution and pathogens.

09 BIOMASS FUELS↗
DOE OSTI2020

Reducing host DNA contamination in 16S rRNA gene surveys of anthozoan microbiomes using PNA clamps

Efforts to study the microbial communities associated with corals can be limited by inefficiencies in the sequencing process due to high levels of host amplification by universal bacterial 16S rRNA gene primers. Here, we develop an inexpensive peptide nucleic acid (PNA) clamp that binds to a target sequence of host DNA during PCR and blocks amplification. We then test the ability of this PNA clamp to mitigate host contamination and increase overall microbial sequence coverage on samples from three coral species: the gorgonians Eunicea flexuosa and Gorgonia ventalina, and the scleractinian Porites panamensis. The 20-bp PNA clamp was designed using DNA from E. flexuosa. Adding the PNA clamp during PCR increased the percentage of microbial reads in E. flexuosa samples more than 11-fold. Microbial community diversity was similar without- and with-PNA clamps, as were the relative frequencies of the ten most abundant ASVs (amplicon sequence variants), indicating that the clamps successfully blocked host DNA amplification while simultaneously increasing microbial DNA amplification proportionally across the most abundant taxa. The reduction of E. flexuosa DNA correlated with an increase in the abundance of rarer taxa. The clamp also increased the average percentage of microbial reads in another gorgonian, G. ventalina, by 8.6-fold without altering the microbial community beta diversity, and in a distantly related scleractinian coral, P. panamensis, by nearly double. The reduction of host contamination correlated with the number of nucleotide mismatches between the host amplicon and the PNA clamp. The PNA clamp costs as little as $0.48 per sample, making it an efficient and cost-effective solution to increase microbial sequence coverage for high-throughput sequencing of coral microbial communities.

16S rRNA gene sequencing↗
DOE OSTI2018

Electron-Impact Excitation of Uracil Luminescence on a Ceramic Surface

Photoelectron spectroscopy was applied to pyrimidine nitrogenous bases, an important class of six-membered heterocyclic compounds incorporated into nucleic acids. The emission spectrum of uracil adsorbed on a ceramic surface that was obtained by bombardment with 600-eV electrons in a high vacuum was analyzed. Broad bands with maxima at 335, 435, and 495 nm were observed in the UV and visible regions. The strongest band (λ = 335 nm) was attributed to fluorescence and corresponded to a singlet–singlet transition from the first excited electronic state into the molecular ground state. Electronic transitions from a triplet T{sub 1} into the ground state formed a weaker phosphorescence band (λ = 435 nm). The nature of the band maximum at 495 nm is discussed. The obtained luminescence spectrum was compared with photoluminescence spectra in various phases.

74 ATOMIC AND MOLECULAR PHYSICS↗
DOE OSTI2018

Use of Fourier Transform IR Spectroscopy for the Study of Saliva Composition

The IR spectrum of saliva may be divided into groups of absorption bands corresponding to lipids, proteins, nucleic acids, and sugars. The finding of thiocyanate ion absorption bands is a characteristic feature of the spectra of saliva. In comparing a typical IR spectrum of saliva with the spectra of albumin, glucose, and lysozyme, a similarity is noted at 1100–1040 cm{sup –1} with the spectrum of albumin and at 1700–1400 cm{sup –1} with the spectra of glucose and lysozyme. It was shown that sugars are found in saliva in a form bound with proteins. Correlations were found between band intensities in the IR spectrum of saliva and its biochemical composition.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2017

Analytical Techniques Used to Detect DNA Binding Modes of Ruthenium(II) Complexes with Extended Phenanthroline Ring

This review describes the analytical techniques used to detect DNA-probes such as Ru(II) complexes with hetero cyclic imidazo phenanthroline (IP) ligands. Studies on drug-DNA interactions are useful biochemical techniques for visualization of DNA both in vitro and in vivo. The interactions of small molecules that binds to DNA are mainly classified into two major classes, one involving covalent binding and another non-covalent binding. Covalent binding in DNA can be irreversible and may leads to inhibition of all DNA processes which subsequently leads to cell death. Usually, covalent interactions leads to permanent changes in the structure of nucleic acids. The non-covalent interaction of molecules with DNA can be due to electrostatic interaction, intercalation and groove binding. These interactions of DNA probes can be explored by various spectroscopic techniques viz. UV–visible, emission, emission quenching spectroscopy, viscosity and thermal denaturation measurements.

74 ATOMIC AND MOLECULAR PHYSICS↗
DOE OSTI2022

Electron microscopy holdings of the Protein Data Bank: the impact of the resolution revolution, new validation tools, and implications for the future

Abstract As a discipline, structural biology has been transformed by the three-dimensional electron microscopy (3DEM) “Resolution Revolution” made possible by convergence of robust cryo-preservation of vitrified biological materials, sample handling systems, and measurement stages operating a liquid nitrogen temperature, improvements in electron optics that preserve phase information at the atomic level, direct electron detectors (DEDs), high-speed computing with graphics processing units, and rapid advances in data acquisition and processing software. 3DEM structure information (atomic coordinates and related metadata) are archived in the open-access Protein Data Bank (PDB), which currently holds more than 11,000 3DEM structures of proteins and nucleic acids, and their complexes with one another and small-molecule ligands (~ 6% of the archive). Underlying experimental data (3DEM density maps and related metadata) are stored in the Electron Microscopy Data Bank (EMDB), which currently holds more than 21,000 3DEM density maps. After describing the history of the PDB and the Worldwide Protein Data Bank (wwPDB) partnership, which jointly manages both the PDB and EMDB archives, this review examines the origins of the resolution revolution and analyzes its impact on structural biology viewed through the lens of PDB holdings. Six areas of focus exemplifying the impact of 3DEM across the biosciences are discussed in detail (icosahedral viruses, ribosomes, integral membrane proteins, SARS-CoV-2 spike proteins, cryogenic electron tomography, and integrative structure determination combining 3DEM with complementary biophysical measurement techniques), followed by a review of 3DEM structure validation by the wwPDB that underscores the importance of community engagement.

Burley, Stephen K. (ORCID:0000000224879713)↗
DOE OSTI2021

Photosynthesis | Photosynthetic Carbon Dioxide Fixation

Carbon dioxide is reduced to make the molecules of life such as proteins, lipids, carbohydrates, and nucleic acids and as the primary mechanism of energy storage for the biosphere. Reduction of fully oxidized carbon, carbon dioxide, occurs following carboxylation of preexisting molecules rather than direct reduction of carbon dioxide to formic acid or formaldehyde. Furthermore, the primary pathway is called the Calvin Benson cycle and proceeds mostly as a series of gluconeogenic (reversal of glycolytic) reactions plus a few pentose phosphate pathway reactions.

Aldolase reaction↗
DOE OSTI2015

Virology: The Next Generation from Digital PCR to Single Virion Genomics

In the past 25 years, virology has had major technology breakthroughs stemming first from the introduction of nucleic acid amplification testing, but more recently from the use of next-generation sequencing, digital PCR, and the possibility of single virion genomics. These technologies have and will improve diagnosis and disease state monitoring in clinical settings, aid in environmental monitoring, and reveal the vast genetic potential of viruses. Using the principle of limiting dilution, digital PCR amplifies single molecules of DNA in highly partitioned endpoint reactions and reads each of those reactions as either positive or negative based on the presence or absence of target fluorophore. In this review, digital PCR will be highlighted along with current studies, advantages/disadvantages, and future perspectives with regard to digital PCR, viral load testing, and the possibility of single virion genomics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Development and characterization of a hydrogen peroxide-resistant cholangiocyte cell line: A novel model of oxidative stress-related cholangiocarcinoma genesis

Oxidative stress is a cause of inflammation–related diseases, including cancers. Cholangiocarcinoma is a liver cancer with bile duct epithelial cell phenotypes. Our previous studies in animal and human models indicated that oxidative stress is a major cause of cholangiocarcinoma development. Hydrogen peroxide (H{sub 2}O{sub 2}) can generate hydroxyl radicals, which damage lipids, proteins, and nucleic acids, leading to cell death. However, some cells can survive by adapting to oxidative stress conditions, and selective clonal expansion of these resistant cells would be involved in oxidative stress-related carcinogenesis. The present study aimed to establish H{sub 2}O{sub 2}-resistant cell line from an immortal cholangiocyte cell line (MMNK1) by chronic treatment with low-concentration H{sub 2}O{sub 2} (25 μM). After 72 days of induction, H{sub 2}O{sub 2}-resistant cell lines (ox-MMNK1-L) were obtained. The ox-MMNK1-L cell line showed H{sub 2}O{sub 2}-resistant properties, increasing the expression of the anti-oxidant genes catalase (CAT), superoxide dismutase-1 (SOD1), superoxide dismutase-2 (SOD2), and superoxide dismutase-3 (SOD3) and the enzyme activities of CAT and intracellular SODs. Furthermore, the resistant cells showed increased expression levels of an epigenetics-related gene, DNA methyltransferase-1 (DNMT1), when compared to the parental cells. Interestingly, the ox-MMNK1-L cell line had a significantly higher cell proliferation rate than the MMNK1 normal cell line. Moreover, ox-MMNK1-L cells showed pseudopodia formation and the loss of cell-to-cell adhesion (multi-layers) under additional oxidative stress (100 μM H{sub 2}O{sub 2}). These findings suggest that H{sub 2}O{sub 2}-resistant cells can be used as a model of oxidative stress-related cholangiocarcinoma genesis through molecular changes such as alteration of gene expression and epigenetic changes. - Highlights: • An H{sub 2}O{sub 2}-resistant ox-MMNK1-L cells was established from immortalized cholangiocytes. • The resistance was acquired by daily treatment of low H{sub 2}O{sub 2} (25 μM) for 15 passages. • The cells highly expressed catalase, SODs and DNMT1 with rapid cell proliferation. • Pseudopodia and the loss of cell-to-cell adhesion appeared by 100 μM H{sub 2}O{sub 2} treatment. • The resistant cells can be used as a model of oxidative stress-related carcinogenesis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

High inorganic phosphate causes DNMT1 phosphorylation and subsequent fibrotic fibroblast activation

Phosphate is an essential constituent of critical cellular functions including energy metabolism, nucleic acid synthesis and phosphorylation-dependent cell signaling. Increased plasma phosphate levels are an independent risk factor for lowered life-expectancy as well as for heart and kidney failure. Nevertheless, direct cellular effects of elevated phosphate concentrations within the microenvironment are poorly understood and have been largely neglected in favor of phosphor-regulatory hormones. Because interstitial fibrosis is the common determinant of chronic progressive kidney disease, and because fibroblasts are major mediators of fibrogenesis, we here explored the effect of high extracellular phosphate levels on renal fibroblasts. We demonstrate that high inorganic phosphate directly induces fibrotic fibroblast activation associated with increased proliferative activity, increased expression of α-smooth muscle actin and increased synthesis of type I collagen. We further demonstrate that such fibroblast activation is dependent on phosphate influx, aberrant phosphorylation of DNA methyltransferase DNMT1 and aberrant CpG island promoter methylation. In summary, our studies demonstrate that elevated phosphate concentrations induce pro-fibrotic fibroblast activation independent of phospho-regulatory hormones. - Highlights: • We exposed human kidney fibroblasts to media containing 1 mM or 3 mM phosphate. • Increased phosphate influx causes phosphorylation of DNA methyltransferase Dnmt1. • Phosphorylated Dnmt1 causes promoter methylation and transcriptional silencing of RASAL1. • Depletion of RASAL1 causes increased intrinsic Ras-GTP activity and fibroblast activation. • Inorganic phosphate causes fibroblast activation independent of phospho-regulatory hormones.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

A novel role for the nuclear localization signal in regulating hnRNP K protein stability in vivo

hnRNP K is a highly conserved nucleocytoplasmic shuttling protein, which associates with RNAs through synergistic binding via its three KH domains. hnRNP K is required for proper nuclear export and translational control of its mRNA targets, and these processes are controlled by hnRNP K's movement between subcellular compartments. Whereas the nuclear export and localization of hnRNP K that is associated with mRNP complexes has been well studied, the trafficking of hnRNP K that is unbound to mRNA has yet to be elucidated. To that end, we expressed an EGFP-tagged RNA binding-defective form of hnRNP K in intact Xenopus embryos, and found it was rapidly degraded in vivo. Deleting hnRNP K's nuclear localization signal (NLS), which contains two prospective ubiquitination sites, rescued the protein from degradation. These data demonstrate a novel activity for the NLS of hnRNP K in regulating the protein's stability in vivo when it is unbound to nucleic acids. - Highlights: • An RNA-binding mutant of Xenopus hnRNP K is degraded in vivo. • hnRNP K's nuclear localization signal contains two predicted ubiquitination sites. • Deletion of the nuclear localization signal rescues RNA-binding mutant degradation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Increased cellular uptake of peptide-modified PEGylated gold nanoparticles

Gold nanoparticles are promising drug delivery vehicles for nucleic acids, small molecules, and proteins, allowing various modifications on the particle surface. However, the instability and low bioavailability of gold nanoparticles compromise their clinical application. Here, we functionalized gold nanoparticles with CPP fragments (CALNNPFVYLI, CALRRRRRRRR) through sulfhydryl PEG to increase their stability and bioavailability. The resulting gold nanoparticles were characterized with transmission electron microscopy (TEM), dynamic light scattering (DLS), UV-visible spectrometry and X-ray photoelectron spectroscopy (XPS), and the stability in biological solutions was evaluated. Comparing to PEGylated gold nanoparticles, CPP (CALNNPFVYLI, CALRRRRRRRR)-modified gold nanoparticles showed 46 folds increase in cellular uptake in A549 and B16 cell lines, as evidenced by the inductively coupled plasma atomic emission spectroscopy (ICP-AES). The interactions between gold nanoparticles and liposomes indicated CPP-modified gold nanoparticles bind to cell membrane more effectively than PEGylated gold nanoparticles. Surface plasmon resonance (SPR) was used to measure interactions between nanoparticles and the membrane. TEM and uptake inhibitor experiments indicated that the cellular entry of gold nanoparticles was mediated by clathrin and macropinocytosis. Other energy independent endocytosis pathways were also identified. Our work revealed a new strategy to modify gold nanoparticles with CPP and illustrated the cellular uptake pathway of CPP-modified gold nanoparticles.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Thermostable DNA helicase improves the sensitivity of digital PCR

Highlights: • A thermostable helicase (Tk-Upf1) unwound duplex DNA with a 5′ to 3′ polarity. • Tk-Upf1 eliminated noise DNAs derived from primer mis-annealing in PCR. • The thermostable helicase improved sensitivity of SNPs detection in digital PCR. DNA/RNA helicases, which catalyze the unwinding of duplex nucleic acids using the energy of ATP hydrolysis, contribute to various biological functions involving DNA or RNA. Euryarchaeota-specific helicase Tk-EshA (superfamily 2) from the hyperthermophilic archaeon Thermococcus kodakarensis has been used to decrease generation of mis-amplified products (noise DNAs) during PCR. In this study, we focused on another type (superfamily 1B) of helicase, Tk-Upf1 (TK0178) from T. kodakarensis, and compared its effectiveness in PCR and digital PCR with that of Tk-EshA. For this purpose, we obtained Tk-Upf1 as a recombinant protein and assessed its enzymatic characteristics. Among various double-stranded DNA (dsDNA) substrates (forked, 5′ overhung, 3′ overhung, and blunt-ended duplex), Tk-Upf1 had the highest unwinding activity toward 5′ overhung DNAs. Noise DNAs were also eliminated in the presence of Tk-Upf1 at concentrations 10-fold lower than those required to yield a comparable reduction with Tk-EshA. When a 5′ or 3′ overhung mis-annealed primer was included as a competitive primer along with specific primers, noise DNAs derived from the mis-annealed primer were eliminated in the presence of Tk-Upf1. In digital PCR, addition of Tk-EshA or Tk-Upf1 increased fluorescent intensities and improved separation between common and risk allele clusters, indicating that both helicases functioned as signal enhancers. In comparison with Tk-EshA, a smaller amount of Tk-Upf1 was required to improve the performance of digital PCR.

60 APPLIED LIFE SCIENCES↗