General Information
The seed of this species was purchased from Chris Greenwell from Bulgaria. Exact flower color is not known but most populations range from pink to purple. Some populations have albino forms. Named for green stripes on two of the sepals. This species is found on grassy meadows on limestone soils.
Plating Media: F522 medium + 2 uM MetaTopolin + 4, 1 cm3 cubes Russet potato sliced into quarters, 25 mins. 0.525% Chlorox bleaching or F522 medium + 2 uM MetaTopolin + 4, 1 cm3 cubes Russet potato sliced in quarters + 20 ml/l fresh pineapple juice. 25 mins 0.525% chlorox bleaching. The media containing the pineapple juice was adjusted to a pH of 5.6. The other media was not adjusted or checked. The seeds were plated on 150 mm diameter petri dishes containing 100 ml of plating media. Plates were sealed with parafilm and incubated in the dark.
Both platings were initiated on 1/8/2012. The plates were checked on 2/26/12 and both media revealed excellent percent germination and very fast germination. There was quite a size range of protocorms.
Number of Days between Plating and Replating: approximately 48 days
Replating Media: Two replating media were used:
1) F522 medium + 1, 1 cm3 cube Russet potato sliced in quarters + 20 ml/l pineapple juice + .5g/l fine charcoal + 1.5g/l culture gel
2) F522 medium + 1, 1 cm3 cube Russet potato sliced in quarters + 20 ml/l pineapple juice + no charcoal + 1.5 g/l culture gel
Both media were adjusted approximately to pH 5.6. The protocorms were replated into wide mouth pint jars containing 200 ml of media, 20 protocorms per pint jar. Because there was considerable variation in protocorm size on the original plates, the protocorms were replated on two different dates. On media 1 above 120 of the larger protocorms were replated on 2/29/2012. On media 2 above 250 slower growing protocorms were replated on 4/24/2012. Replates were incubated in the dark at room temperature ( probably 22-26 o C).
Replates on media 1 were placed in the frig on Aug. 4 after 159 days of growth. Replates on media 2 were placed in the frig on Aug. 17 after only 121. The only real difference between the two media was the presence of charcoal. The difference in results were striking. Plants grown without charcoal (media 2) had taller shoots and more roots than those growing on media 1 and in some cases the shoots reached the top of the pint jars. However the roots of plants in media 2 although significant in length showed the release of brown phenolic compounds. Plants grown with charcoal had shorter shoots ranging in height from a couple of millimeters to about 1.5 inches. The roots of plants grown in charcoal were either absent or much shorter but looked better-- no phenolic compounds visible. After vernalization and growing them at room temperature under lights can the better media be ascertained.
Number of Days between Replating and Beginning of Vernalization: 159 days with media 1 and 121 days in media 2.
Number of Days between start and end of Vernalization: 120 days (4 months)
Date to Initiate Plating: Jan.1
Date to Replate: Feb. 29 approximately
Date to start to Vernalize: August 4-August 17
Final Results after Vernalization and growth under lights: