Journal
/
Genetics
/
Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions
JoVE Journal
Genetics
A subscription to JoVE is required to view this content.  Sign in or start your free trial.
JoVE Journal Genetics
Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions

Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions

DOI:
10.3791/58051-v

•

08:23 min

•

September 25, 2018

•

, , , , , ,
1Circulating Tumor Biomarkers Laboratory, SiRIC, Translational Research Department,Institut Curie, PSL Research University, 2CNRS UMR144,Institut Curie, PSL Research University, 3Department of Medical Oncology,Institut Curie, PSL Research University, 4University Versailles Saint-Quentin-en-Yvelines, 5University Paris Descartes, 6Inserm U830,Institut Curie, PSL Research University

Chapters

  • 00:04Title
  • 00:48Blood Collection and Plasma Storage
  • 02:04cfDNA Extraction
  • 04:45ddPCR Mix Preparation
  • 05:46Droplet Generation
  • 06:25PCR Amplification
  • 07:06Results: KRAS and EGFR Drop-off ddPCR Assays
  • 07:50Conclusion

Summary

Automatic Translation

Automatic Translation

Here, we present a protocol to accurately quantify multiple genetic alterations of a target region in a single reaction using drop-off ddPCR and a unique pair of hydrolysis probes.

Tags

Single Droplet Digital PCRCirculating Tumor DNAMutation DetectionHotspot RegionsCancer DiagnosisAbsolute QuantificationMolecular AnalysisQPCR AlternativePlasma ExtractionProteinase KLysis BufferSilica MembraneVacuum Pump

Article

Embed

ADD TO PLAYLIST

Usage Statistics

Related Videos

Improved Polymerase Chain Reaction-restriction Fragment Length Polymorphism Genotyping of Toxic Pufferfish by Liquid Chromatography/Mass Spectrometry

The Visual Colorimetric Detection of Multi-nucleotide Polymorphisms on a Pneumatic Droplet Manipulation Platform

Wild-type Blocking PCR Combined with Direct Sequencing as a Highly Sensitive Method for Detection of Low-Frequency Somatic Mutations

Detection of Copy Number Alterations Using Single Cell Sequencing

Retroviral Scanning: Mapping MLV Integration Sites to Define Cell-specific Regulatory Regions

Semi-quantitative Detection of RNA-dependent RNA Polymerase Activity of Human Telomerase Reverse Transcriptase Protein

CRISPR-Mediated Reorganization of Chromatin Loop Structure

Differentiation, Maintenance, and Analysis of Human Retinal Pigment Epithelium Cells: A Disease-in-a-dish Model for BEST1 Mutations

Identification of Homologous Recombination Events in Mouse Embryonic Stem Cells Using Southern Blotting and Polymerase Chain Reaction

Evaluation of a Universal Nested Reverse Transcription Polymerase Chain Reaction for the Detection of Lyssaviruses

Read Article