INVITRO ASSAY OF EXCISED COTYLEDONS OF ALFALFA (MEDICAGO-SATIVA) TO SCREEN FOR RESISTANCE TO COLLETOTRICHUM-TRIFOLII

An in vitro assay using excised cotyledons of alfalfa has been developed that accurately detects resistance to Colletotrichum trifoli. The adaxial surfaces excised cotyledons are placed on sterile moist filter paper in petri plates. The cotyledons are sprayed with a spore suspension, and resistant and susceptible reactions can be determined after incubation for 14 days at 24 C with a 16-hr light period. Susceptibility is based on the presence of sporulating acervuli. Five isolates of C. trifolii reacted similarly in the assay, and the range of effective inoculum was 2,500-10,000 spores per spray. Excised cotyledons and the seedlings from which the cotyledons were obtained were simultaneously tested in vitro and in a greenhouse. Saranac AR and Saranac had 86.5 and 96.2% agreement of observations, respectively, between the two screening techniques. A blind test of six cultivars was done, and the percent resistance obtained with the in vitro screen agreed with published values. Results indicate that usefulness of this technique as an alternative to greenhouse screening methods.