Persivore

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  1. Persivore

    Inermis seeds germinating

    I used to use bleach (calcium hypochlorite) for seed sterilization, but have moved away from it since PPM seems to be much more effective. Bleach would be successful about 90% of the time, whereas PPM has been 100% successful (so far). Strangely, i found N.inermis seed to be the most difficult to sterilize - I went through a few failed batches of these, sterilizing with bleach, before moving to PPM. If you catch the contamination early enough you can usually save most of the batch by picking out the affected seeds, re-sterilizing and re-plating. I tend to move any seedlings which germinate to a separate container once they are free of the seed casing, in case contamination hits. These are my sterilization methods if you get a chance to give tissue culture another go - Calcium hypochlorite Seeds are soaked in 250 parts per million gibberellic acid/distilled water for 24 hours. I typically use about 20ml of this 250ppm GA3 solution. The GA3 itself doesn't sterilize anything, but the soaking probably softens up the seeds a bit and helps to bring any fungus spores out of dormancy to make them easier to kill. After 24 hours, I add 100mg to 150mg of calcium hypochlorite powder to this 20ml to give 0.5-0.75% bleach solution. This is shaken until most of the calcium hypochlorite is dissolved. After 10-15 minutes, the seeds are rinsed in sterile distilled water 3 times and moved to culture jars. Household bleach can be used as an alternative to calcium hypochlorite powder, but the quantities will need adjusting based on the bleach strength. PPM Seeds are again soaked in 250ppm GA3/water solution for 24 hours. After 24 hours the GA3 solution is drained and a 1:10 PPM:water solution (1ml PPM + 10ml DI water) is added to the seeds and left for a further 24 hours. The PPM solution is then drained and the seeds moved to culture jars without rinsing. The PPM solution can be re-used if kept refrigerated. Keep up the good work - you'll soon have more Nepenthes than you can cope with!
  2. Persivore

    Inermis seeds germinating

    Try sterilizing your seeds by soaking them in a 1:10 mix of PPM and water (1ml ppm and 10ml DI water). I let the nepenthes seeds soak in this PPM solution for 24 hours, then drain the PPM off and place the seeds on culture media containing 1ml/litre of PPM. I've had zero contamination problems since moving to this method. The PPM sterilization solution can be reused i you keep it refrigerated. Germinating Nepenthes seeds in tissue culture really is worth the extra effort, once you sort out any contamination problems.