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.
|Dettol hand sanitiser|
|Holstered auto-pipette and pipette box|
Now purchase a bag of eppendorf tubes from the same lab consumables supplier (see pic). There are various volumes to choose from but get the graduated 1500µl or 2000µl tubes with lockable caps. Your next investment will be a vortex mixer (see pic) which will ensure that the seeds will be continuously aggitated and mixed with the sterilising agent for the correct amount of time and will avoid dead-spots associated with other techniques. Vortexing in the eppendorfs also concentrates the seeds nicely allowing better control for rinsing and pouring of seeds onto plates. Eppendorfs are also autoclavable and I place a fistful into a polypropylene tub with a sealable lid dry into the pressure cooker for sterilising. Once sterile, I remove this tub of eppendorfs to the laminar flow. You can include the vortex mixer inside the laminar flow if you have the space but you can also have it next to the hood on the working bench if you wish. In the hood you should also have a sterile flask of RO water and a flask of pre-made sodium hypochlorite solution to which some surfactant has been added. Use the following concentration of non-perfumed liquid sodium hypochlorite: 120ml tap water to 20ml neat household sodium hypochlorite solution (3.5% m/v). Add a few drops of Tween20 or household dish-washing liquid (not too much or it will foam too much!). The surfactant facilitates good soaking by allowing the sodium hypochlorite solution to "stick" to the surface of the seeds.
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.