PCR amplify the ITS region, a fungal DNA barcode that you can sequence and match to genetic databases for molecular identification of fungi.

• Techniques: Micropipetting, DNA extraction, PCR, gel electrophoresis, bioinformatics
• Topics: Biodiversity monitoring, DNA barcoding, species identification
• Time required: This is a project-based activity that requires at least three class periods for completion
• Level: Advanced high school, college, independent research

This kit contains the materials needed to uniquely identify fungal species using DNA technology. There are hundreds of thousands of known species of wild mushrooms and other fungi in the world. And millions more that we are yet to discover! Fungi can be found in virtually all known ecosystems, yet many species look similar to each other, making them difficult to identify by eye.

Start with your own samples, from the mold on your overripe tomato to wild-harvested fruiting mushrooms. Once you have extracted their DNA, you will be able to amplify 80 unique fungal DNA barcodes by PCR and, with the help of an add-on kit and a sequencing service, obtain DNA sequences for up to 40 of them.
The Fungal Add-On Kit contains primers, nuclease-free water, and tubes aimed to facilitate submission of amplified ITS PCR products for Sanger Sequencing.

Reagents and consumables supplied by user:
- Agarose (electrophoresis grade)
- DNA stain (e.g., SeeGreen™ or GelGreen®)
- Gel electrophoresis buffer (e.g., 1X TBE)
- Distilled or deionized H₂O (to dilute buffer concentrate – not included in Lab Companion Kit)
- Microtubes
- PCR tubes
- Fungal samples

Required equipment:
- Micropipettes and tips
- Tissue homogeneizer or vortex mixer with microtube holder attachment
- Microcentrifuge
- Thermal cycler
- Gel electrophoresis and visualization system
- Sanger DNA sequencing service

Storage notes:
- Reagents may ship at room temperature, but require freezer storage upon arrival
- Frozen reagents are stable for 12 months when stored in freezer