A entomopathogenic ascomycetous fungus, Cordyceps militaris is distributed world wide and cultivated as a medicinal mushroom having many valuable biological and pharmaceutical activities. In the breeding of new C. militaris mushroom, single ascospores were isolated and tested their characteristics such as mycelial growth, mycelial density, fruiting body formation ability, the production of perithecia. Also two opposite mating types were assayed using two sets of primers specific for C. militaris, which were amplified a 191-bp fragment for MAT1-1 and 233-bp fragment for MAT1-1. F1 hybrids resulted in well-developed perithecial fruiting bodies and their crossings were confirmed by the PCR assays. New hybrids were produced showing high quality fruiting bodies in artificial media. Mycelial growth of new strain 'Dowonhongcho 2’ was better on SDAY and at 15-25°C when it was compared with that of 'Yedang 3'. The stromata of new strain were club-shaped and bright orange-red color. Its length was 7.1 cm and the cordycepin content was 0.33% on average. In comparison with 'Yedang 3', the new strain had a yield that was 7% higher and it produced fruiting bodies which were firmer. Days of fruiting body were similar with 50 days from inoculation. This cultivar may serve as a valuable one for artificial cultivation and industrial-scale production of C. militaris.

For heaths and vitality Cordyceps sinensis has been widely used as an important medicinal mushroom. C. militaris is popular for its substitute and can be cultivated artificially. Its new strains have been improved by crossing single spore strains. A bipolar heterothallic fungus, C. militaris has two strains of compatible mating types and they can be differentiated by crossing, fruiting body formation ability and the production of perithecia. Each isolate was determined to be monokaryotic or dikaryotic on the basis of perithecial formation by fruiting tests. This process was also carried out by molecular markers for the detection of two opposite mating types or F1 hybrids using two sets of primers specific for C. militaris amplifying a 191-bp fragment for MAT1-2 and 233-bp fragment for MAT1-1. Therefore the crossing of two compatible mating types resulted in well-developed perithecial fruiting bodies and their crossings were confirmed by the PCR assays for the rapid and specific detection of both MAT1-1 and MAT1-2. This results showed that molecular markers provide an efficient alternative for identification of hybridal status or mating types in the breeding program for artificial cultivation and industrial-scale production of C. militaris.

Cordyceps species are important mushrooms traditionally used for heaths and vitality. C. sinensis has been widely used as a medicinal mushroom and C. militaris is popular for its substitute. C. militaris can be cultivated artificially and new strains has been improved by crossing single spore strains. As a bipolar heterothallic fungus C. militaris has two strains of compatible mating types and they can be differentiated by crossing, fruiting body formation ability and the production of perithecia. However this process is very laborious and time consuming to carry out. In this study, molecular markers were designed and used for the detection of two opposite mating types. Two mating types were assayed using two sets of primers specific for C. militaris, which were amplified a 191-bp fragment for MAT1-2 and 233-bp fragment for MAT1-1. After crossing of two compatible mating types F1 hybrids resulted in well-developed perithecial fruiting bodies and their crossings were confirmed by the multiplex PCR assays for the rapid and specific detection of both MAT1-1 and MAT1-2. This results may serve as a efficient process in the breeding program for artificial cultivation and industrial-scale production of C. militaris.

Single ascospores were isolated and tested their characteristics such as mycelial growth, mycelial density, fruiting body formation ability, the production of perithecia in the breeding of new Cordyceps militaris mushroom. Among them selected isolates were crossed and hybrids were produced showing high quality fruiting bodies in artificial media. New strain 'Dowonhongcho’ was better on SDAY and at 10~25°C when it was compared with that of 'Yedang 3' in mycelial growth. The stromata of new strain were club-shaped and bright orange-red color. Its length was 6.1 cm and the cordycepin content was 0.34% on average. In comparison with 'Yedang 3', the new strain had a yield that was 9% higher and it produced fruiting bodies which were firmer. The optimum temperature for mycelial growth was 22~25°C and the optimum temperature for stroma development was 18~22°C. Days of fruiting body were similar with 45 days from inoculation. This cultivar may serve as a valuable one for artificial cultivation and industrial-scale production of C. militaris.