Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Method for preparing single-stranded DNA templates for Pyrosequencing using vector ligation and universal

Marco Groth1, Klaus Huse, Kathrin Reichwald

  • 1Genome Analysis, Leibniz Institute for Age Research-Fritz Lipmann Institute, 07745 Jena, Germany. mgroth@fli-leibniz.de

Analytical Biochemistry
|June 2, 2006
PubMed
Summary

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Estimation of controlled attenuation parameter-based liver steatosis via raw ultrasound data from handheld devices.

Journal of medical imaging (Bellingham, Wash.)·2026
Same author

Ingestible active capsule for gastrointestinal microbiome sampling.

Biomedical microdevices·2026
Same author

Current Trends in Ultrasound Wearables: Spotlight on System Architecture.

IEEE reviews in biomedical engineering·2026
Same author

<i>In vivo</i> evaluation of a biodegradable intraanastomotic membrane in a porcine model.

Frontiers in surgery·2026
Same author

Evaluation of image quality in five different handheld ultrasound devices and analysis of various impact factors.

Ultraschall in der Medizin (Stuttgart, Germany : 1980)·2026
Same author

Real-Time In Vivo Monitoring of Anastomotic Intestinal Ischemia Using Implantable Resorbable Organic Sensors.

Advanced science (Weinheim, Baden-Wurttemberg, Germany)·2025

This study introduces a new method for Pyrosequencing DNA template preparation, reducing the need for expensive primers. The novel approach simplifies single-stranded DNA template isolation for genetic analysis.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Pyrosequencing is a valuable technique for DNA sequencing, providing quantitative and qualitative data.
  • Current Pyrosequencing methods require single-stranded DNA templates, typically generated using PCR with specific biotinylated primers.
  • The need for numerous expensive, template-specific biotinylated primers presents a significant cost and logistical challenge.

Purpose of the Study:

  • To develop a cost-effective and streamlined method for preparing single-stranded DNA templates for Pyrosequencing.
  • To overcome the limitations associated with using numerous template-specific biotinylated primers in DNA template preparation.

Main Methods:

  • A novel method involving the ligation of amplified DNA fragments into a plasmid vector was developed.

Related Experiment Videos

  • This facilitated subsequent PCR using a universal, vector-specific biotinylated primer.
  • The method enables straightforward isolation of single-stranded templates from any PCR product.

    • Main Results:

    • The developed method successfully simplifies the isolation of single-stranded DNA templates.
    • Proof-of-principle experiments demonstrated its utility in genotyping single-nucleotide polymorphisms (SNPs) in CARD15 and A2M genes.
    • The method was also effective for characterizing multisite variations in the DEFB104 gene.

    Conclusions:

    • The new ligation-based approach offers an efficient and economical alternative for preparing single-stranded DNA templates for Pyrosequencing.
    • This method significantly reduces the reliance on expensive, custom-synthesized biotinylated primers.
    • The technique is broadly applicable for genetic analysis, including SNP genotyping and variation characterization.