I received a few inquires about how I germinated D.Regia seeds on compost. This was my first attempt at growing what I affectionately call the "uglies". They are by no means a plant I like, I am just an equal opportunity germinator. I read a couple of threads on other's experiences germinating and growing D.regia from seed. The take home message: 1) the seed had better be viable at the start and 2) feed the babies or they will suffer the consequences. Below, is a more detailed protocol of what I did. For those of you who I had previously responded to, please go by the notes below. After having sowed hundreds of containers in the last few months, I can't recall on the fly which babies got rolled in what. 
Soil
I used a 2:1:1 mix of peat:sand: pumice.
- Sand = HTH pool filter sand, 20 grit, 50 lb bag. If you use a lot of sand, this is very economical. I was buying 5lb bags of horticulural grade sand at OSH but this works just as well. I pour some in a 4L beaker and fill it up with milliQ water. I will mix the sand and change the water until is is clear, and no longer milky.
- Peat = Black gold peat. This has saponin as a wetting agent. I don't wash this - that is where I draw the line. ( FYI, saponin is a detergent (amphiphatic) and is used on various grains like quinoa and should be washed if one is going to consume whatever it is covering. It imparts a bitter taste.)
- Pumice = Black gold pumice (Yes, that is all they have at my local OSH). This stuff is VERY dirty (twigs/rocks/ probably my grandma if I looked hard enough..etc) and necessitates multiple washes/rinses.
The ingredients are mixed in the ratio above and should be moist, not soggy. It is placed in a 2L glass beaker, covered with foil and autoclaved for 20-25 minutes. Usually I make this ahead of time and have some on hand because it needs to cool to room temperature before use. Excess soil can be stored in the 22C.
Seed Pretreatment
In a small petri dish microwave some water to lukewarm temperature. It should be cool enough to touch but still warm. Add a "sprinkle" of trichoderma flowerable (Ampac Biotech, store 4C) - sorry that is what my notes say. Mix the trichoderma solution (color is light brown) and add the seeds. Allow the seeds to swim in the trichoderma mix at room temperature for 1 hour (at least).
Sowing Seeds
I sowed the pretreated seeds in a 4'' round plastic pot, the diameter which is perfect for making a dixie cup greenhouse. This is my emulation from RL7836 posts on the ICPS. I thought it would be a cheap way so I could watch them grow, as I am not a big fan of the ziplock baggies. (1) 2mm hole was made in the center top of the dixie cup with a surgical knife. In another clean glass beaker, I added the necessary volume of cool autoclaved soil mix and ~1/40th of a tsp of granular trichoderma (Ampac Biotech). The soil was not filled to the brim of the pot, ~1/2'' from the brim so the dixie cup could effectually "sit on top of the soil" snugly. Using a loop, I fished out each seed from the trichoderma brew and placed it onto the soil. I gave each seed a gentle "poke" to make sure it is snug in its new home. The seeds were evenly spaced from each other in the pot, all 1'' away from edge of the pot. I had already planned on dividing each little sprout into its own pot shortly after germination. This space between the seeds was to allow me to scoop each seedling + soil as a plug, and place it into a new pot without disturbing them (who likes earthquakes?). After all was snug, I placed the dixie cup over it, said a prayer, and into the incubator it went.
Germination Temperature, Lighting, and Watering
If you have read my other thread "Nepenthes Tissue Culture", you may remember that I have turned my planaria incubator into my germination chamber. (This is only for the interim until the planaria reach larger numbers - when I will have to move my plants elsewhere. Good thing one of my postdocs could not perform molarity calculations, or maybe not..).
Temperature -The incubator temperature is set to 21C, 24 hours a day, 365 days a year. The temperature inside the dixie cup is 76-78F high/69F low.
Light - Light cycle is 16hr ON/ 8 hr OFF. Light is provided by: 8watt, 320 lumens 13’’ F8T5CW 4200K + 24'' 20watt Phillips Daylight 6500K. The distance of the brim of the pot to the light bulb = 4''.
Watering - I mist with autoclaved milliQ h20 (I happened to have a lot on hand usually since we use a lot in the lab) every day or every other day. One sprout did not make it past sprouting and the verdict is still out if this was a result of desiccation, or if the seed was simply not up to snuff.
Seedling Temperature, Lighting, Feeding, Watering, and Transplanting
Temperature - The "uglies" have not seemed to fret over the intermediate conditions I have been providing them. Seedlings, with at least 2 true leaves, like to be closer, rather than farther, to the light provided that one can keep the temperature below 80F. There have been days when the incubator has been open for extended amounts of time and the temperature was at the low 80'sF.
Lighting -The 1'' "uglies" are separated onto 2 different shelves. One set is 9'' from the light and the other is ~3'' from light. Those 3'' from the light were 1.5X the size of those 9'' from the light at 5 weeks old. Yesterday, I moved those 9'' from the light closer to the 3'' by elevating them on empty pipet tip boxes.
Feeding - At 2 true leaves, the "uglies" are still tiny and I didn't expect much root formation so I placed the slow release oscomote pellets around the seedlings. The pellets were close, 1/2'' from base of each the "uglies". Basically, they were surrounded by at least 2 pellets each. This is critical. After having to starve oneself for many years to make weight on weigh in days, it makes one very unhappy and metabolic stress pathways turn on in full force. Feed them!
Watering - I HATE WATERING!! And even so, I must mist every 2 days at least. The dixie cups allow a lot of moisture to escape since the soil is not completely level, so they are checked on periodically. However, I don't water the plants. Instead, I give each oscomote pellet a good misting.
Transplanting - Each "ugly" was given its own 4'' pot + dixie cup when they started bumping into the sides of the dixie cup and I saw that the hairy real root started coming off its backside (4 true leaves at this point). I thought it prudent to move them before the root rooted. Using my trusty spoon (not the same one I eat with) which has been autoclaved, I spooned out each "ugly" + plug-of-soil into a new pot with same soil mix used originally. A little depression the size of the plug has been carved out to facilitate the process. Level the ground and surround the "ugly" with 5 or so, oscomote pellets. The pellets were placed a little farther this time, 1'' , to encourage the roots to grow out.
"Uglies" got 2 true leaves so get their first feeding of oscomote pellets
First week after oscomote pellets
"Uglies" hairy backside -> time to transplant
"Uglies" in their new homes with more pellets
Incubator Growing Chamber - Second level is where D.Regia germinated
Top level -> elevated the "Uglies" growing with empty pipet boxes
Second level
N. Attenboroughii sprout being dwarfed by its "ugly" neighbor
I sowed 11 seeds on compost. 7 germinated. Lost 1. 6 "Uglies" at the end of the day.
Hope this helps people in their endeavors.
Jen
Soil
I used a 2:1:1 mix of peat:sand: pumice.
- Sand = HTH pool filter sand, 20 grit, 50 lb bag. If you use a lot of sand, this is very economical. I was buying 5lb bags of horticulural grade sand at OSH but this works just as well. I pour some in a 4L beaker and fill it up with milliQ water. I will mix the sand and change the water until is is clear, and no longer milky.
- Peat = Black gold peat. This has saponin as a wetting agent. I don't wash this - that is where I draw the line. ( FYI, saponin is a detergent (amphiphatic) and is used on various grains like quinoa and should be washed if one is going to consume whatever it is covering. It imparts a bitter taste.)
- Pumice = Black gold pumice (Yes, that is all they have at my local OSH). This stuff is VERY dirty (twigs/rocks/ probably my grandma if I looked hard enough..etc) and necessitates multiple washes/rinses.
The ingredients are mixed in the ratio above and should be moist, not soggy. It is placed in a 2L glass beaker, covered with foil and autoclaved for 20-25 minutes. Usually I make this ahead of time and have some on hand because it needs to cool to room temperature before use. Excess soil can be stored in the 22C.
Seed Pretreatment
In a small petri dish microwave some water to lukewarm temperature. It should be cool enough to touch but still warm. Add a "sprinkle" of trichoderma flowerable (Ampac Biotech, store 4C) - sorry that is what my notes say. Mix the trichoderma solution (color is light brown) and add the seeds. Allow the seeds to swim in the trichoderma mix at room temperature for 1 hour (at least).
Sowing Seeds
I sowed the pretreated seeds in a 4'' round plastic pot, the diameter which is perfect for making a dixie cup greenhouse. This is my emulation from RL7836 posts on the ICPS. I thought it would be a cheap way so I could watch them grow, as I am not a big fan of the ziplock baggies. (1) 2mm hole was made in the center top of the dixie cup with a surgical knife. In another clean glass beaker, I added the necessary volume of cool autoclaved soil mix and ~1/40th of a tsp of granular trichoderma (Ampac Biotech). The soil was not filled to the brim of the pot, ~1/2'' from the brim so the dixie cup could effectually "sit on top of the soil" snugly. Using a loop, I fished out each seed from the trichoderma brew and placed it onto the soil. I gave each seed a gentle "poke" to make sure it is snug in its new home. The seeds were evenly spaced from each other in the pot, all 1'' away from edge of the pot. I had already planned on dividing each little sprout into its own pot shortly after germination. This space between the seeds was to allow me to scoop each seedling + soil as a plug, and place it into a new pot without disturbing them (who likes earthquakes?). After all was snug, I placed the dixie cup over it, said a prayer, and into the incubator it went.
Germination Temperature, Lighting, and Watering
If you have read my other thread "Nepenthes Tissue Culture", you may remember that I have turned my planaria incubator into my germination chamber. (This is only for the interim until the planaria reach larger numbers - when I will have to move my plants elsewhere. Good thing one of my postdocs could not perform molarity calculations, or maybe not..).
Temperature -The incubator temperature is set to 21C, 24 hours a day, 365 days a year. The temperature inside the dixie cup is 76-78F high/69F low.
Light - Light cycle is 16hr ON/ 8 hr OFF. Light is provided by: 8watt, 320 lumens 13’’ F8T5CW 4200K + 24'' 20watt Phillips Daylight 6500K. The distance of the brim of the pot to the light bulb = 4''.
Watering - I mist with autoclaved milliQ h20 (I happened to have a lot on hand usually since we use a lot in the lab) every day or every other day. One sprout did not make it past sprouting and the verdict is still out if this was a result of desiccation, or if the seed was simply not up to snuff.
Seedling Temperature, Lighting, Feeding, Watering, and Transplanting
Temperature - The "uglies" have not seemed to fret over the intermediate conditions I have been providing them. Seedlings, with at least 2 true leaves, like to be closer, rather than farther, to the light provided that one can keep the temperature below 80F. There have been days when the incubator has been open for extended amounts of time and the temperature was at the low 80'sF.
Lighting -The 1'' "uglies" are separated onto 2 different shelves. One set is 9'' from the light and the other is ~3'' from light. Those 3'' from the light were 1.5X the size of those 9'' from the light at 5 weeks old. Yesterday, I moved those 9'' from the light closer to the 3'' by elevating them on empty pipet tip boxes.
Feeding - At 2 true leaves, the "uglies" are still tiny and I didn't expect much root formation so I placed the slow release oscomote pellets around the seedlings. The pellets were close, 1/2'' from base of each the "uglies". Basically, they were surrounded by at least 2 pellets each. This is critical. After having to starve oneself for many years to make weight on weigh in days, it makes one very unhappy and metabolic stress pathways turn on in full force. Feed them!
Watering - I HATE WATERING!! And even so, I must mist every 2 days at least. The dixie cups allow a lot of moisture to escape since the soil is not completely level, so they are checked on periodically. However, I don't water the plants. Instead, I give each oscomote pellet a good misting.
Transplanting - Each "ugly" was given its own 4'' pot + dixie cup when they started bumping into the sides of the dixie cup and I saw that the hairy real root started coming off its backside (4 true leaves at this point). I thought it prudent to move them before the root rooted. Using my trusty spoon (not the same one I eat with) which has been autoclaved, I spooned out each "ugly" + plug-of-soil into a new pot with same soil mix used originally. A little depression the size of the plug has been carved out to facilitate the process. Level the ground and surround the "ugly" with 5 or so, oscomote pellets. The pellets were placed a little farther this time, 1'' , to encourage the roots to grow out.
"Uglies" got 2 true leaves so get their first feeding of oscomote pellets
First week after oscomote pellets
"Uglies" hairy backside -> time to transplant
"Uglies" in their new homes with more pellets
Incubator Growing Chamber - Second level is where D.Regia germinated
Top level -> elevated the "Uglies" growing with empty pipet boxes
Second level
N. Attenboroughii sprout being dwarfed by its "ugly" neighbor
I sowed 11 seeds on compost. 7 germinated. Lost 1. 6 "Uglies" at the end of the day.
Hope this helps people in their endeavors.
Jen
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