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Related Concept Videos

Next-generation Sequencing03:00

Next-generation Sequencing

The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.
Sanger Sequencing01:57

Sanger Sequencing

DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
RNA-seq03:21

RNA-seq

RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...
The Replisome03:01

The Replisome

DNA replication is carried out by a large complex of proteins that act in a coordinated matter to achieve high-fidelity DNA replication. Together this complex is known as the DNA replication machinery or the replisome.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with the...
Lagging Strand Synthesis01:59

Lagging Strand Synthesis

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.
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...
Real Time RT-PCR02:57

Real Time RT-PCR

Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...

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Related Experiment Video

Updated: Jun 27, 2026

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes
05:37

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes

Published on: April 4, 2025

Real-time DNA sequencing from single polymerase molecules.

John Eid1, Adrian Fehr, Jeremy Gray

  • 1Pacific Biosciences, 1505 Adams Drive, Menlo Park, CA 94025, USA.

Science (New York, N.Y.)
|November 22, 2008
PubMed
Summary
This summary is machine-generated.

This study demonstrates real-time DNA sequencing using fluorescently labeled nucleotides and nanostructures. The method achieves high accuracy, revealing polymerase dynamics and DNA secondary structures during synthesis.

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Direct Observation of Enzymes Replicating DNA Using a Single-molecule DNA Stretching Assay
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Last Updated: Jun 27, 2026

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes
05:37

Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes

Published on: April 4, 2025

DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis
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DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis

Published on: October 6, 2017

Direct Observation of Enzymes Replicating DNA Using a Single-molecule DNA Stretching Assay
17:03

Direct Observation of Enzymes Replicating DNA Using a Single-molecule DNA Stretching Assay

Published on: March 23, 2010

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Genomics

Background:

  • Accurate DNA sequencing is crucial for understanding biological processes and disease.
  • Previous methods faced limitations in real-time observation and accuracy.

Purpose of the Study:

  • To develop a single-molecule, real-time DNA sequencing method.
  • To analyze DNA polymerase dynamics and identify sequence-dependent features.

Main Methods:

  • Utilized zero-mode waveguide nanostructure arrays for parallel single-molecule detection.
  • Employed fluorescently labeled deoxyribonucleoside triphosphates (dNTPs) conjugated to terminal phosphates.
  • Monitored enzymatic incorporation in real-time over thousands of bases.

Main Results:

  • Achieved continuous observation of DNA synthesis without steric hindrance.
  • Identified distinct polymerase states and pause sites linked to DNA secondary structures.
  • Generated consensus sequences with a median accuracy of 99.3%.

Conclusions:

  • This novel sequencing approach provides direct insights into polymerase kinetics.
  • The method accurately determines DNA sequences and reveals biophysical parameters of polymerization.
  • Demonstrated high accuracy and potential for identifying sequence-specific errors.