CLASSICAL METHODS AND SPECIFIC PRIMERS IN DETECTION OF ENDOPHYTIC FUNGI IN SOME GRAMINEUOS PLANTS

Mutual relationship between endophytic fungi and some gramineuos plants is one of the recently known host-microbe interactions. Endophytic fungi are classified in class Ascomycetes, order Hypocreales, tribe Balansieae and mostly genus Neotyphodium. They are transferred into the progeny via maternal stock and have many benificial effects to their host plants, such as resistance to pests and diseases, environmental stresses such as pH fluctuation, drought, salinity and grazing. because Iran has a wide range of plant genomic resources and is known as one of the origins of gramineous plants, it is necessary to know more about endophytic fungi and their presence by using classical and molecular methods. Twenty isolates of endophytic fungi were obtained from seed and leaf sheaths of Bromus tomentellus, Lolium prenne, Festuca arundinacea and Festuca pratensis. Mycelia of endophytic fungi were observed red or pink in seed and leaf sheath when stained by Rose Bengal. The DNA of endophytic fungi was extracted by Murry and Thompson method with minor modification. In order to confirm the identity of endophytes, three universal and specific primer e. g. ITS1, ITS4, IS1, IS3, and 11-1, 11-2 were used. All isolates showed a 550-750 bp band with ITS1, ITS4 and eighteen of them showed a 444 bp band with IS1, IS3 primers, which is an indicator of Neotyphodium endophytes. when primers 11-1, 11-2 were used, a 1000 bp band was not observed in all isolates. Isolates FaSm and FpGon3 that did not have shown a 444 bp band, may probably differ from Neotyphodium endophytic fungi.