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Southern Blot

Southern Blot

Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments.
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...

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Updated: Jun 27, 2026

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

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

Split hybridisation probes for electrochemical typing of single-nucleotide polymorphisms.

Fausto Lucarelli¹, Silvia Capponcelli, Giovanna Marrazza

¹Department of Chemistry, University of Florence, via della Lastruccia 3, 50019 Sesto F.no, Florence, Italy.

|December 17, 2008

Summary

This study introduces a novel method for highly selective single-nucleotide polymorphism (SNP) typing using split hybridization probes. The assay accurately detects single-base mutations at room temperature, proving effective for clinical genetic analysis.

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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

Published on: September 25, 2018

Area of Science:

Molecular Biology
Biosensing Technology
Genetics

Background:

Accurate detection of single-nucleotide polymorphisms (SNPs) is crucial for genetic analysis and disease diagnosis.
Existing SNP typing methods often require strict temperature control and can lack specificity.
Developing a robust, room-temperature SNP detection method is highly desirable for clinical applications.

Purpose of the Study:

To develop and validate a highly selective SNP typing method utilizing split hybridization probes.
To demonstrate the electrochemical analysis of single-base mutations in patient samples using this novel assay.
To provide simple rules for optimizing probe design for high specificity and mismatch discrimination.

Main Methods:

Development of a SNP typing assay based on split hybridization probes and allele-specific oligonucleotide (ASO) binding.
Electrochemical analysis using disposable probe-modified gold electrodes as the genosensing platform.
Application of the assay to detect mutations in synthetic targets and clinical samples (TP53 gene), including those within hairpin structures, using auxiliary oligonucleotides.

Main Results:

The split hybridization probe assay demonstrated high selectivity and full mismatch discrimination at ambient conditions.
Nanomolar detection limits were achieved for synthetic targets in the presence of excess mismatched sequences.
The assay successfully discriminated single-base mutations in clinical TP53 gene targets, with detection limits in the high picomolar range (60 min total analysis time).

Conclusions:

The developed split hybridization probe assay offers a simple, cost-effective, and highly selective method for SNP typing.
The assay is suitable for low-throughput analysis of clinically relevant samples, even for challenging targets.
This approach provides a robust platform for electrochemical genosensing with potential for broader diagnostic applications.