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Related Concept Videos

Single-Strand DNA Binding Proteins01:03

Single-Strand DNA Binding Proteins

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For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
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During replication, the complementary strands in double-stranded DNA are synthesized at different rates. Replication first begins on the leading strand. Replication starts later, occurs more slowly, and proceeds discontinuously on the lagging strand.
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DNA from cells is required for many biotechnology and research applications, such as molecular cloning. To remove and purify DNA from cells, researchers use various methods of DNA extraction. While the specifics of different protocols may vary, some general concepts underlie the process of DNA extraction.
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DNA isolation protocols can be fast and straightforward or complex and time-consuming depending on the type and quality of DNA required for further processing. For example, plasmid DNA extraction is a bit more complicated than genomic DNA extraction because of the need for an appropriate lysis method to separate plasmid DNA from gDNA during isolation. However, for specific applications, such as long-range DNA sequencing that require a good yield of high- quality DNA samples, we need to follow...
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Fixing Double-strand Breaks02:04

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The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
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A Droplet-Based Microfluidic Approach and Microsphere-PCR Amplification for Single-Stranded DNA Amplicons
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Isolating single stranded DNA using a microfluidic dialysis device.

Yixiao Sheng1, Michael T Bowser

  • 1Department of Biomedical Engineering, University of Minnesota, Minneapolis, MN 55455, USA.

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|November 12, 2013
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Summary
This summary is machine-generated.

This study introduces a microfluidic device for separating single-stranded DNA (ssDNA) from double-stranded DNA (dsDNA). The novel device achieves high purity ssDNA, crucial for biotechnology applications like aptamer generation.

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Microfluidics

Background:

  • Isolating single-stranded DNA (ssDNA) from double-stranded DNA (dsDNA) is vital for aptamer generation and other biotech applications.
  • Existing methods can be cumbersome and less efficient for large-scale applications.

Purpose of the Study:

  • To develop and validate a microfluidic, flow-through dialysis device for efficient ssDNA isolation from dsDNA.
  • To optimize the device's performance by evaluating the impact of NaOH concentration and flow rate.

Main Methods:

  • Fabrication of a two-channel polydimethylsiloxane (PDMS) microfluidic device separated by a polycarbonate membrane.
  • Immobilization of dual-biotinylated dsDNA onto streptavidin-coated beads.
  • Alkaline denaturation to release ssDNA, followed by membrane separation in a flow-through dialysis system.

Main Results:

  • Achieved >95% ssDNA purity using 25 mM NaOH.
  • Identified that lower flow rates are necessary to maximize ssDNA yield, approaching the theoretical 50% maximum.
  • The microfluidic device demonstrated superior ssDNA purity compared to manual methods under optimized conditions.

Conclusions:

  • The developed microfluidic device offers a highly efficient and pure method for isolating ssDNA.
  • This technology has significant potential for advancing aptamer generation and other molecular biology techniques.
  • Optimization of NaOH concentration and flow rate is key to balancing purity and yield.