Browsing by Author "Chen, Wen-Yih"

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Improved DNA detection by utilizing electrically neutral DNA probe in field-effect transistor measurements as evidenced by surface plasmon resonance imaging
(2013-03-15)
Chen, Wen-Yih
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Chen, Hon-Chen
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Yang, Yuh-Shyong
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Huang, Chun-Jen
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Chan, Hardy Wai-Hong
Intensive efforts have been focused on the development of ultrasensitive DNA biosensors capable of quantitative gene expression analysis. Various neutralized nucleic acids have been demonstrated as alternative and attractive probe for the design of a DNA chip. However, the mechanism of the improvements has not been clearly revealed. In this investigation, we used a newly developed neutral ethylated DNA (E-DNA), a DNA analog with the "RO-P-O" backbone (wherein R could be methyl, ethyl, aryl, or alkyl group) obtained from synthetic procedures, and a silicon nanowire (SiNW) field-effect transistor (FET) to evaluate the difference in DNA detection performance while using E-DNA and DNA as probes. It is demonstrated that using the E-DNA probe in the FET measurement could have a significantly enhanced effect upon the detection sensitivity. Surface plasmon resonance imaging (SPRi) was used to evidence the mechanism of the improved detection sensitivity. SPRi analysis showed the amounts of probe immobilization on the sensor surface and the hybridization efficiency were both enhanced with the use of E-DNA. Consequently, neutral ethylated DNA probe hold a great promise for DNA sensing, especially in the electrical-based sensor. (C) 2012 Elsevier B.V. All rights reserved.
Neutralized chimeric DNA probe for detection of single nucleotide polymorphism on surface plasmon resonance biosensor
(2018-01-15)
Huang, Chun-Jen
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Lin, Zhong-En
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Yang, Yuh-Shyong
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Chan, Hardy Wai-Hong
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Chen, Wen-Yih
An implementation of neutralized chimeric DNA oligomer as a probe for sensitive detection of single nucleotide polymorphisms (SNPs) on a surface plasmon resonance imaging sensor is investigated. The chimeric DNA oligomer was synthesized in a conventional DNA synthesizer, containing neutral nucleotides with a methylated phosphate group. The secondary structures and melting points of the chimeric DNA fragment and its complexes with perfect-matched and single-mismatched complementary DNA molecules were examined by using circular dichroism and UV vis spectroscopy in comparison with the native probe DNA counterpart. The results indicate that the chimeric DNA complexes can form a B-form structure and exhibit high thermostability. Moreover, the hybridization and discrimination efficiency of the chimeric probe DNA for the SNP genotyping were verified by using the SPRi sensor under different experimental conditions. The data reveal the effects of the ionic strength and operation temperature on the selectivity of the chimeric probe DNA for the SNP detection. The hybridization condition with a low ionic strength and high temperature allows the chimeric probe DNA distinguishing perfect-matched and single-mismatched target DNA molecules to the best extent, likely due to the reduced electrostatic repulsive force and presence of the additional methyl group on the backbone. Consequently, the direct and label-free detection with the SPR technique and neutralized chimeric probe DNA can be realized for the SNP genotyping by optimizing the operation condition and sequence design.
Neutralized chimeric DNA probe for the improvement of GC-rich RNA detection specificity on the nanowire field-effect transistor
(2019-07-30)
Chou, Wei-Cheng
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Hu, Wen-Pin
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Yang, Yuh-Shyong
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Chan, Hardy Wai-Hong
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Chen, Wen-Yih
Silicon nanowire (SiNW) field-effect transistors (FETs) is a powerful tool in genetic molecule analysis because of their high sensitivity, short detection time, and label-free detection. In nucleic acid detection, GC-rich nucleic acid sequences form self-and cross-dimers and stem-loop structures, which can easily obtain data containing signals from nonspecific DNA binding. The features of GC-rich nucleic acid sequences cause inaccuracies in nucleic acid detection and hinder the development of precision medicine. To improve the inaccurate detection results, we used phosphate-methylated (neutral) nucleotides to synthesize the neutralized chimeric DNA oligomer probe. The probe fragment originated from a primer for the detection of hepatitis C virus (HCV) genotype 3b, and single-mismatched and perfect-matched targets were designed for single nucleotide polymorphisms (SNP) detection on the SiNW FET device. Experimental results revealed that the HCV-3b chimeric neutralized DNA (nDNA) probe exhibited better performance for SNP discrimination in 10 mM bis-tris propane buffer at 25 degrees C than a regular DNA probe. The SNP discrimination of the nDNA probe could be further improved at 40 degrees C on the FET device. Consequently, the neutralized chimeric DNA probe could successfully distinguish SNP in the detection of GC-rich target sequences under optimal operating conditions on the SiNW FET device.
PEGylation Site-Dependent Structural Heterogeneity Study of MonoPEGylated Human Parathyroid Hormone Fragment hPTH(1-34)
(2014-09-30)
Liu, Chih-Ying
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Li, Xin
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Chen, Wen-Yih
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Chang, Li-Chiao
;
Chen, Yi-Fan
The structures of C- and N-terminally monoPEGylated human parathyroid hormone fragment hPTH(1-34) as well as their unmodified counterparts, poly(ethylene glycol) (PEG) and hPTH(1-34), have been studied by small-angle neutron scattering (SANS). The scattering results show that free hPTH(1-34) in 100 mM phosphate buffer (pH 7.4) aggregates into clusters. After conjugation with PEG, the PEG-peptide conjugates self-assemble into a supramolecular core-shell structure with a cylindrical shape. The PEG chains form a shell around the hPTH(1-34) core to shield hPTH(1-34) from the solvent. The detailed structural information on the self-assembled structures is extracted from SANS using a model of the cylindrical core with a shell of Gaussian chains attached to the core surface. On the basis of the data, because of the charge-dipole interactions between the conjugated PEG chain and the peptide, the conjugated PEG chain forms a more collapsed conformation compared to free PEG. Moreover, the size of the self-assembled structures formed by the C-terminally monoPEGylated hPTH(1-34) is about 3 times larger than that of the N-terminally monoPEGylated hPTH(1-34). The different aggregation numbers of the self-assembled structures, triggered by different PEGylation sites, are reported. These size discrepancies because of different PEGylation sites could potentially affect the pharmacokinetics of the hPTH(1-34) drug.
Reduction of interstrand charge repulsion of DNA duplexes by salts and by neutral phosphotriesters - Contrary effects for harnessing duplex formation
(2020-05-01)
Kuo, Tai-Chih
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Wu, Meng-Wei
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Lin, Wei-Chen
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Matulis, Daumantas
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Yang, Yuh-Shyong
To exploit the formation of DNA duplexes, the charge repulsion between complementary DNA strands can be reduced by adding salts or by replacing the negatively charged natural phosphodiester linkage( s) with charge-neutral methyl phosphotriester(s) (MPTE) linkage( s) of the synthetic nDNA oligonucleotides. Recently, we reported prominent improvements of detecting target nucleic acids with nDNA probes on liquid-phase PCR and solid/liquid-interface biosensing. To understand the observed improvements from thermodynamics perspectives, we studied the formation and stability of the double-stranded DNA (dsDNA) containing nDNA and/or natural DNA. With the natural DNA oligonucleotides, increment of Na+ cations caused unexpected reductions of favorable enthaply (Delta H) of duplex formation and expected increases of favorable free energy (Delta G) and temperature of melting transition (T-m). With one nDNA in duplex formation, the increase of Na+ yielded extra faborable Delta H, yet unfavorable Delta G and T-m . Possibly, Na(+ )cations ( > 50 mM) promote the formation of dsDNA through the endothermic release of DNA (and nDNA)-hydrating water molecules more than through the reduction of inter-strand repulsions. The methyl groups of MPTE of nDNA also destabilize the DNA duplexes by hindering the formation of standard double helices. These novel findings explain the duplex-destabilization property of nDNA which enables its discrimination of single-nucleotide polymorphisms. (C) 2020 Taiwan Institute of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
Signal Enhancement of Silicon Nanowire Field-Effect Transistor Immunosensors by RNA Aptamer
(2019-09-17)
Vu, Cao-An
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Hu, Wen-Pin
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Yang, Yuh-Shyong
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Chan, Hardy Wai-Hong
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Chen, Wen-Yih
Silicon nanowire field-effect transistors (SiNW-FETs) have been demonstrated as a highly sensitive platform for label-free detection of a variety of biological and chemical entities. However, detecting signal from immunoassays by nanoFETs is severely hindered by the distribution of different charged groups of targeted entities, their binding orientation, and distances to the surface of the FET. Aptamers have been widely applied as a recognition element for plentiful biosensors because of small molecular sizes and moderate to high specific binding affinity with different types of molecules. In this study, we propose an effective approach to enhance the electrical responses of both direct (6x-histidine) and sandwich (amyloid beta 1-42) immunoassays in SiNW-FETs with R18, a highly negative charged RNA aptamer against rabbit immunoglobulin G (IgG). Empirical results presented that the immunosensors targeted with R18 expressed a significantly stabilized and amplified signal compared to the ones without this aptamer. The research outcome provides applicability of the highly negative charged aptamer as a bioamplifier for immunoassays by FETs.
Synergetic improvements of sensitivity and specificity of nanowire field effect transistor gene chip by designing neutralized DNA as probe
(2018-08-22)
Hu, Wen-Pin
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Tsai, Chih-Chin
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Yang, Yuh-Shyong
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Chan, Hardy Wai-Hong
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Chen, Wen-Yih
Neutral DNA analogs as probes for the detection of target oligomers on the biosensors based on the field-effect transistor (FET) configuration feature advantages in the enhancement of sensitivity and signal-to-noise ratio. Herein, we used phosphate-methylated nucleotides to synthesize two partially neutralized chimeric DNA products and a fully neutralized DNA sequence and adopted a regular DNA oligomer as probes on the polycrystalline silicon nanowire (NW) FET devices. The sequences of two neutralized chimeric DNAs close to the 5' end were alternately modified with the phosphate-methylated nucleotides, and all probes were immobilized via their 5' end on the NW surface. The non-specific-to-specific binding ratio indicated that the two 5'-end partially neutralized chimeric DNAs featured better performance than the regular and fully neutralized DNA oligomers. The partially neutralized probe design reduces the ionic strength needed for hybridization and increases the Debye length of detection, thus promoting the detection sensitivity of FET and achieving the limit of detection of 0.1 fM. By using an appropriate probe design, applying DNA oligomers with embedded phosphate-methylated nucleotides in the FET biosensors is a promising way for gene detection with high sensitivity and specificity.
Synergy of using neutralized DNA on nanowire field effect transistor as next generation biosensor
(2016-07-25)
Chen, Wen-Yih
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Yang, Yuh-Shyong
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Chan, Hardy W.
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交大名義發表
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National Chiao Tung University
Synergy of using neutralized DNA on nanowire field effect transistor as next generation biosensor
(2016-07-25)
Chen, Wen-Yih
;
Yang, Yuh-Shyong
;
Chan, Hardy W.
;
交大名義發表
;
National Chiao Tung University