Counting DNA molecules on a microchannel surface for quantitative analysis

Kim, Taesoo; Kim, Siwon; Noh, Chanyoung; Hwang, Hyeseung; Shin, Jieun; Won, Nabin; Lee, Seonghyun; Kim, Dogeun; Jang, Yoon-ha; Hong, Su-Jin; Park, Jungyul; Kim, Sung Jae; Jang, Seongsoo; Lim, Kwang‐il; Jo, Kyubong

doi:10.1016/j.talanta.2022.123826

Cited by 5 publications

(5 citation statements)

References 35 publications

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“…The semiconductive properties of silicon wafers effectively mitigate charging effects, [ 19 ] and the silicon oxide layer can be derivatized using quaternary ammonium siloxane to immobilize the DNA backbone. [ 10 ] To explore the influence of surface attributes, we applied a droplet of PVP solution to four different surfaces. After drying, the edges of the droplets were captured using SEM.…”

Section: Resultsmentioning

confidence: 99%

“…Preparation of Positively Charged Silicon Wafer Surface: Silicon wafers with a 30-nm SiO 2 layer were purchased from Wafer Market (Yong-In, Korea). The method established for glass coverslips [10] was adapted to treat the SiO 2 layer and create positively charged surfaces. The wafers were incubated in a 250-mL solution containing N-trimethoxysilylpropyl-N, N, Ntrimethylammonium chloride (1.1 mm) at 65 °C with agitation at 100 rpm for 16 h. They were then rinsed three times with 99.9% ethanol and stored in 99.9% ethanol.…”

Section: Dna-binding Protein (Mneongreen-hmg)mentioning

confidence: 99%

“…For example, DNA images obtained using fluorescence microscopy often display well‐aligned and parallel elongations. [ 10 ] This feature is challenging to achieve with TEM, as DNA typically appears randomly arranged due to alignment difficulties on a carbon film‐coated metal grid.…”

Section: Introductionmentioning

confidence: 99%

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Scanning Electron Microscopy Imaging of Large DNA Molecules Using a Metal‐Free Electro‐Stain Composed of DNA‐Binding Proteins and Synthetic Polymers

Noh,

Kang,

Heo

et al. 2024

Advanced Science

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This paper presents the first scanning electron microscopy (SEM)‐based DNA imaging in biological samples. This novel approach incorporates a metal‐free electro‐stain reagent, formulated by combining DNA‐binding proteins and synthetic polymers to enhance the visibility of 2‐nm‐thick DNA under SEM. Notably, DNA molecules stain with proteins and polymers appear as dark lines under SEM. The resulting DNA images exhibit a thickness of 15.0±4.0 nm. As SEM is the primary platform, it integrates seamlessly with various chemically functionalized large surfaces with the aid of microfluidic devices. The approach allows high‐resolution imaging of various DNA structures including linear, circular, single‐stranded DNA and RNA, originating from nuclear and mitochondrial genomes. Furthermore, quantum dots are successfully visualized as bright labels that are sequence‐specifically incorporated into DNA molecules, which highlights the potential for SEM‐based optical DNA mapping. In conclusion, DNA imaging using SEM with the novel electro‐stain offers electron microscopic resolution with the ease of optical microscopy.

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Section: Resultsmentioning

confidence: 99%

Section: Dna-binding Protein (Mneongreen-hmg)mentioning

confidence: 99%

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Scanning Electron Microscopy Imaging of Large DNA Molecules Using a Metal‐Free Electro‐Stain Composed of DNA‐Binding Proteins and Synthetic Polymers

Noh,

Kang,

Heo

et al. 2024

Advanced Science

Self Cite

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show abstract

“…PDMS devices and positively charged surfaces were prepared as previously described . In particular, microchannel templates had two layers on a silicon wafer.…”

Section: Methodsmentioning

confidence: 99%

“…PDMS devices and positively charged surfaces were prepared as previously described. 42 In particular, microchannel templates had two layers on a silicon wafer. First, the SU-82015 photoresist was spin-coated to form a 20 μm photoresist layer with subsequent exposure to 350 nm radiation in a mask aligner (Midas System MDA-400LJ, Daejeon, Korea).…”

Section: ■ Introductionmentioning

confidence: 99%

Twelve Colors of Streptavidin–Fluorescent Proteins (SA-FPs): A Versatile Tool to Visualize Genetic Information in Single-Molecule DNA

et al. 2022

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Streptavidin–fluorescent proteins (SA-FPs) are a versatile tool to visualize a broad range of biochemical applications on a fluorescence microscope. Although the avidin–biotin interaction is widely used, the use of SA-FPs has not been applied to single-molecule DNA visualization. Here, we constructed 12 bright SA-FPs for DNA staining or labeling reagents. To date, 810 FPs are available, many of which are brighter than organic dyes. In this study, 12 bright FPs were selected to construct SA-FP plasmids covering green to red colors. Their brightness ranges from 40 to 165 mM–1 cm–1. Moreover, SA-FP is brighter than FP itself because streptavidin forms a tetramer complex; thus, four FPs are in a single complex. In addition, FPs often form a dimer or a tetramer, resulting in multiple FPs in a single spot on a microscopic image. This feature is advantageous because multiple fluorescent β-barrels on a single biotin tag provide enough brightness to be easily visualized by epifluorescence microscopy. Using SA-FPs, we visualized DNA backbones, nickase-based optical mapping, and AT-frequency profiling. Finally, we demonstrated the combination of nickase-based optical mapping using SA-FP and AT-frequency profiling.

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Microfluidic Device to Maximize Capillary Force Driven Flows for Quantitative Single-Molecule DNA Analysis

Kim

2023

BioChip J

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Counting DNA molecules on a microchannel surface for quantitative analysis

Cited by 5 publications

References 35 publications

Scanning Electron Microscopy Imaging of Large DNA Molecules Using a Metal‐Free Electro‐Stain Composed of DNA‐Binding Proteins and Synthetic Polymers

Scanning Electron Microscopy Imaging of Large DNA Molecules Using a Metal‐Free Electro‐Stain Composed of DNA‐Binding Proteins and Synthetic Polymers

Twelve Colors of Streptavidin–Fluorescent Proteins (SA-FPs): A Versatile Tool to Visualize Genetic Information in Single-Molecule DNA

Microfluidic Device to Maximize Capillary Force Driven Flows for Quantitative Single-Molecule DNA Analysis

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