Aug 08, 2018
Version 2
- 1Laboratorio GeMBio, Centro de Investigacion Cientifica de Yucatan A.C
External link: 10.1007/s10658-012-0065-7
Protocol Citation: Daisy Perez-Brito 2018. ISOLATION OF PHYTOPATHOGENIC FUNGI. protocols.io https://dx.doi.org/10.17504/protocols.io.sgsebwe
Manuscript citation:
Rodríguez-García CM, Ruiz-Ruiz JC, Peraza-Echeverría L, Peraza-Sánchez SR, Torres-Tapia LW, Pérez-Brito D, Tapia-Tussell R, Herrera-Chalé FG, Segura-Campos MR, Quijano-Ramayo A, Ramón-Sierra JM, Ortiz-Vázquez E (2019) Antioxidant, antihypertensive, anti-hyperglycemic, and antimicrobial activity of aqueous extracts from twelve native plants of the Yucatan coast. PLoS ONE 14(3): e0213493. doi: 10.1371/journal.pone.0213493
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: August 08, 2018
Last Modified: August 08, 2018
Protocol Integer ID: 14578
Keywords: Phytopathogenic fungi, Colletotrichum spp, Alternaria spp
Abstract
To control a disease it is essential to know its causal agent. A correct diagnosis provides a considerable amount of basic information to select correct control methods. An accurate diagnosis always starts with a representative sample of the damaged tissue, from which a portion is taken, to isolate the causal agent. In the case of phytopathogenic fungi, there are several isolation methods, depending on the tissue or substrate in which they are found. This protocol describes the isolation and purification of phytopathogenic fungi from leaves and fruits of different tropical crops.
Attachments
Materials
STEP MATERIALS
3% NaOCl solution
water-agar medium (2%)
3% NaOCl solution
water-agar medium (2%)
Protocol materials
water-agar medium (2%)
3% NaOCl solution
water-agar medium (2%)
3% NaOCl solution
3% NaOCl solution
water-agar medium (2%)
Fungi Isolation
Fungi Isolation
Cut diseased plant tissues, taken from the advanced margin of lesions, into small pieces (5 × 5 mm) with a scalpel.
Disinfest by immersing them in 3% NaOCl solution.
3% NaOCl solution
Rinse with sterile distilled water. (1/3)
Rinse with sterile distilled water. (2/3)
Rinse with sterile distilled water. (3/3)
Transfer each small piece onto 90 x 15 mm Petri Dish containing Potato Dextrose Agar (PDA) medium.
Incubate at room temperature (25°C) for 7 days.
Fungi Purification
Fungi Purification
Cut in the edge of each fungi colony, with a scalpel a mycelial segment of approximately 5 x 5 mm.
Transfer each small mycelial piece onto 90 x 15 mm Petri dish containing water-agar medium (2%).
water-agar medium (2%)
Incubate at room temperature (25°C) for 5 days.
Observe the fungal colony on a stereo microscope and with an insulin syringe needle (29 gauge) cut one hyphae tip.
Transfer hyphae tip onto 90 x 15 mm Petri Dish containing Potato Dextrose Agar (PDA) medium.
Incubate at room temperature (25°C) for 7 days.