I was browsing through the stats on the blog recently to see what subject seems to be the most popular. It appears as though there are a lot of would-be amateur tissue culture hobbiests out there with a keen interest in how to get started but also how to get things done properly with little fuss. I have read some interesting articles on the internet about home tissue culture of orchids and what methods to follow. Most are basic repetition of the blah-blah basics with some useless stuff perpetuated. Some are good though, especially those which dispense with the drivel about useless or over-complicated techniques. One such "technique" is the tea-bag or waterproof paper technique for surface sterilising dry orchid seeds. Equally irritating is the syringe-method which I threw out many moons ago (in fact about a week after I started). So... since I am abviously irritated by things that challenge my impatience I have decided to share some simple techniques with you that actualy work. In fact, so much so that my entire seedling collection is based upon these since most were cultured as dry seeds.
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Dettol hand sanitiser |
In previous and slightly aged posts, I indicated the importance of understanding the concept of sterility. This is a crucially important first step. Sterility does not mean clean or disinfected, it means absolutely sterile - ie - nothing living! Now consider that everything that is not sterile is covered in bacteria and fungal spores and other microscopic organisms that are just waiting for an opportunity to spread and to multiply. Your basic view now should be that of an obsessive compulsive with a phobia of germs! Possible transmission routes include YOU (your skin, hair, breath, clothes etc.), the air, all surfaces, your tools and kit and the orchid seeds themselves. Some of the most stubborn of bacteria I find are those associated with the actual seeds! Ok, so most of you would be aware that absolutely everything would need to be sterilised before success can be achieved with germinating orchid seeds
in vitro. My archived post on building and using my laminar flow hood should provide you with some basic and useful information on building your own hood for orchid seed use. Although you can do seeds in a glove box or sterile still-air cabinet which I also did at the very beginning, you will never achieve the rate of success that you will with a laminar flow hood. It's worth spending the time and the bucks to make it. Mine cost approximately R2000 for all the materials and construction. For the US readers this equates to about $250. All my tools are cleaned before being sterilised in a 10 quart 15/16 bar pressure cooker for 30 minutes. I wrap all my tools in aluminium foil first so that I can remove them without actually touching them and can transfer them to the laminar flow hood where they can be carefully unwrapped under sterile air and placed into a sterile jar ready for use. I also try not to store too much ready-made plates of media in the fridge and prefer to make a batch of new media the same day or the day before and to allow these to set in the laminar flow with the Hepa running until I use them (so plan ahead - it helps avoid unnecesary non-sterile exposure). Wash your hands and forearms well before working in the hood. Use a good non-sticky hand sanitiser like the one made by Dettol (kills 99.9% germs). Use this liberally on your hands, making sure you get it under your nails and remember to remove your rings and watch before working. I routinely wipe down the inside of the hood working chamber with neat sodium hypochlorite solution before commencing with any work. When the hood is not in use I have a hinged door that seals off the front of the chamber to prevent incidental aerial contamination that might threaten the viability of my Hepa.
I do all of my dry seed work as above. I have found it produces superior results and minimises contamination. I hope you found this post useful. I would love to hear your thoughts and your experiences.
just perfect.
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