On Wed, 31 Jan 1996, Edward Gilding wrote:
> Ok tissue culture people. I got the rest of the procedure down (I think)
> and I am wondering if there is any way to reduce the amount of contamination
> from nepenthes explants in stage 1.
>
> Here's the beef of the problem, Nepenthes aren't all that common and cutting
> them up really breaks my heart, however its much easier to take an explant
> than to chop the poor thing into cuttings. Well, I've taken explants from
> numerous Nepenthes on numerous occaisions, and out of the total of 15-17
> explants taken, only about 3-4 did not die from contamination (but later
> perished from the disinfectant itself, turning brown etc.) I used to use
You can avoid the browning of explants by transfering them immediately
after sterilisation in a solution of citric and ascorbinic acid (dont
remember the exact conc. at this moment.)
> Chlorox often, but I noticed that although it really killed germs, it wasn't
> to easy on the explants too, even when they were soaked for only 5 minutes.
I dont know the ingredients of _Clorox_ but it may contain H2O2 that is
very aggressive for plant tissue. Also with _Domestos_ or NaOCl it is
always diluted 1:5.
> The remaining explants that are alive (and a few of the dead ones) have some
> kind of delayed contamination it is obvious that it does come from the
> explant itself, and almost always from the inside of the leaf axil. Now I
Yes, the phloem / xylem of the plants are often inhabited by
microorganisms such as fungi or bacteria. You cant reach them by surface
sterilisation. You may try antibiotics as media additions but this
also works not 100 %.
> tried peeling back the leaf slightly and letting the disinfectant do its
> business, and it worked better to some degree but all of the explants died.
> The explants are 1, 2-4 node cuttings of mature growth. The article I have
> of Nepenthes TC used axenic derived seedings instead of stem explants.
Please tell us the title of this article.
> Nepenthes seed (viable ones anyway) is not easy to come by.
>
> Has anyone used any alternative method with success at disinfecting
> nepenthes with household ingredients, times of soaking, protocol, etc.
I dont know any alternative method yet.
>
> Thank you
> Edward Gilding
> egilding@lava.net
Hope this is not too disappointing.
Yours
Heiko