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Related Concept Videos

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...
Maxam-Gilbert Sequencing01:05

Maxam-Gilbert Sequencing

In the same year as the discovery of the Sanger sequencing method, another group of scientists, Allan Maxam and Walter Gilbert, demonstrated their chemical-cleavage method for DNA sequencing. The Maxam-Gilbert method relies on using different chemicals that can cleave the DNA sequence at specific sites, the separation of resulting DNA fragments of variable size using electrophoresis, and deciphering the DNA sequence from the resulting gel bands.
Challenges of the Maxam-Gilbert Method
The...
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.
Labeling DNA Probes03:31

Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
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...

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Related Experiment Video

Updated: Jul 7, 2026

DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling
08:04

DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling

Published on: October 8, 2019

Dideoxy DNA sequencing with chemiluminescent detection.

C S Martin1

  • 1Tropix, Inc., Bedford, Massachusetts, USA.

Current Protocols in Molecular Biology
|February 12, 2008
PubMed
Summary

Biotinylated primers enable efficient, nonisotopic DNA sequencing using chemiluminescent detection. This method provides high-resolution DNA sequencing ladders via gel electrophoresis and membrane transfer with enzyme-linked detection.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Standard DNA sequencing methods often rely on isotopic detection, which can be cumbersome and pose safety concerns.
  • Nonisotopic detection methods offer advantages in terms of safety, cost-effectiveness, and ease of use.
  • Chemiluminescent detection provides a sensitive and high-resolution alternative for visualizing DNA sequencing results.

Purpose of the Study:

  • To describe an efficient and nonisotopic method for DNA sequencing using chemiluminescent detection.
  • To detail the use of biotinylated primers for enhanced detection in DNA sequencing reactions.
  • To present a protocol for generating high-resolution DNA sequencing ladders.

Main Methods:

  • Utilizing primers labeled with biotin for DNA sequencing reactions.

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Pyrosequencing for Microbial Identification and Characterization
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Pyrosequencing for Microbial Identification and Characterization

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Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis
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Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Published on: July 8, 2025

Related Experiment Videos

Last Updated: Jul 7, 2026

DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling
08:04

DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling

Published on: October 8, 2019

Pyrosequencing for Microbial Identification and Characterization
12:37

Pyrosequencing for Microbial Identification and Characterization

Published on: August 22, 2013

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis
07:10

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Published on: July 8, 2025

  • Separating reaction products via denaturing gel electrophoresis.
  • Transferring separated DNA to a nylon membrane for detection.
  • Employing streptavidin-alkaline phosphatase conjugates for signal amplification.
  • Detecting the signal using a chemiluminescent 1,2-dioxetane substrate and imaging on X-ray film.

    • Main Results:

    • Successful generation of high-resolution DNA sequencing ladders using chemiluminescent detection.
    • Demonstration of efficient and nonisotopic DNA sequencing.
    • Detailed protocols for indirect labeling with streptavidin and biotinylated alkaline phosphatase.
    • Description of detecting other haptens using antibody-alkaline phosphatase conjugates.

    Conclusions:

    • Nonisotopic, chemiluminescent DNA sequencing with biotinylated primers is an efficient and effective method.
    • The described technique yields high-resolution DNA sequencing results.
    • The protocol is adaptable for detecting various haptens, increasing its versatility.