So before I really get going with tissue culturing of hemlocks, I was wondering how much sterilization of explant material I must do? I have read a few articles and some people use a combination of bleach/alcohol. My plan was to soak in 15% bleach for about 15 minutes, rinse with alcohol, and flame off the excess alcohol. Should that do the job? Am I overdoing it?
First of all, which type of explant are you using? Leaves? Seeds?
Secondly, I think 15% bleach is a bit too concentrated... Try something between 2-5%. 15 min is ok.
Also, I would suggest you invert the order of disinfection: you first dip the explant in 70% alcohol for a few seconds (try 30 seconds) and then in 5% bleach for 15 min. After that, you could rinse it in sterile distilled water three times.
Finally, if you have the time and enough material, try experimenting a little bit. See the least amount of time in disinfection and the least concentrated bleach you could use and still get viable and free of fungi and bacteria explants.
I hope it helps!
Use 20% of bleach concentration at 20 minutes. Thank you and good luck.
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