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Aflatoxigenic fungi DNA isolation for preparation of DNA chip.

Autoři: Šnévajsová Petra | Brožková Iveta | Vytřasová Jarmila

Rok: 2006

Druh publikace: ostatní - přednáška nebo poster

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Jazyk	Název	Abstrakt	Klíčová slova
cze	Izolace DNA pro detekci aflatoxinogenních plísní pomocí DNA čipu.	určeno pro seznam publikací
eng	Aflatoxigenic fungi DNA isolation for preparation of DNA chip.	This work covers possibilities of DNA isolation and immobilization from the toxinogenic fungi of genus Aspergillus for preparation of DNA chip. Filamentous fungi have a sturdy cell wall. The most difficult step in the isolation of DNA from Aspergillus species is to disrupt cell wall without causing damage to genomic DNA. Two classical techniques of DNA extraction were optimized and compared with use of commercial PCR kits. Classical techniques included DNA extraction by Cenis (1992) and modified procedure by Shapira et al. (1996) in connection with Cenis (1992). Fungi mycelium was disrupted by ultrasonic bath and liquid nitrogen. Furthermore three commercial kits for DNA isolation from plant leaves, seeds and plants were used. For DNA isolation was also used commercially delivered compound DNAzol?ES. DNA was purified by immobilization on superparamagnetic silica particles and diatom. Methods were optimized by using pure cultures A. parasiticus var. globosus CCM F ? 550 and A. flavus CCM F ? 108. Acquired DNA was amplified by PCR. Specific PCR product (apa-2 gene) was detected by electrophoresis. Records were interpreted. Purity and concentration of DNA were identified by spectrophotometer. Modified procedure by Shapira et al. (1996) in which liquid nitrogen is used to disrupt cell wall has proved as very effective. The PCR product coding biosynthetic way of aflatoxin B1 will use for construction of DNA chip.	DNA chip, isolation, A.flavus

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