Related Experiment Video
Updated: Jun 17, 2026
Isolation, Characterization, and Total DNA Extraction to Identify Endophytic Fungi in Mycoheterotrophic Plants
Published on: May 5, 2023
[Basic methods for extraction and molecular analysis of micromycetic DNA]
Abstract:
The methods for extraction of nucleic acids from the fungi and clinical materials containing various microorganisms have currently modified and make it possible to extract DNA with its highest yield and to minimize its destruction. In particular, this concerns mycelial fungi that have a rigid cell wall. Addition of glass microbeads to the lysed sample, followed by mixture stirring, gives rise to undamaged DNA from fungi and their spores. Extracted DNA is suitable for the identification of micromycetes by qualitative or quantitative polymerase chain reaction (PCR), real-time PCR-based hybridization, and LightCycler PCR and for the polymorphism analysis of the spacer regions of ribosomal genes by the sequencing technique. A combination of quantitative PCR and sequencing permits co-determination of the number of gene target copies in 9 different micromycetes in the clinical material being examined, which is of great importance for the diagnosis and treatment of patients with higher immunosuppression and, of those after transplantation of organs and tissues in particular.
Insights
This study presents an improved DNA extraction method using glass microbeads for fungi, ensuring high yield and integrity. This method enables accurate identification and quantification of fungal pathogens in clinical samples, crucial for immunocompromised patients.
Area of Science:
- Molecular Biology
- Mycology
- Clinical Diagnostics
Context:
- Fungal infections pose significant risks, especially to immunocompromised individuals.
- Accurate identification and quantification of fungal pathogens are critical for effective treatment.
- Traditional DNA extraction methods can struggle with rigid fungal cell walls, leading to low yield and DNA degradation.
Purpose:
- To develop and optimize a DNA extraction method for fungal nucleic acids from clinical samples.
- To ensure high yield and minimize degradation of DNA from various microorganisms, particularly mycelial fungi.
- To validate the suitability of the extracted DNA for molecular diagnostic techniques.
Summary:
- A modified DNA extraction protocol utilizing glass microbeads effectively lyses rigid fungal cell walls, yielding high-quality, intact DNA from fungi and their spores.
- The extracted DNA is suitable for various molecular analyses, including qualitative/quantitative polymerase chain reaction (PCR), real-time PCR, and sequencing for ribosomal gene polymorphism.
- Combining quantitative PCR and sequencing allows for the simultaneous determination of gene target copy numbers for up to nine different micromycetes in clinical specimens.
Impact:
- This method enhances the accuracy and efficiency of diagnosing fungal infections in clinical settings.
- It is particularly beneficial for patients with compromised immune systems, such as organ transplant recipients.
- Improved fungal DNA extraction facilitates precise diagnosis and targeted treatment strategies, improving patient outcomes.
Related Concept Videos
01:24DNA Isolation
01:34DNA Isolation
01:29Modern Molecular Taxonomy