I want to observe inoculated bacteria on rice tissue. Some bacteria were tagged with gfp while other with bfp.
The question is a little bit general, so you should give a little bit more information about some things that can influence the answer. For example, are the bacteria inside the root (or leaf) tissue or outside, on the surface?
If they are inside, you can try with a paraformaldehyde based fixative (with no glutaraldehyde, what produces autofluorescence in plant tissues). Fix the tissues, and then you can cut them by hand, vibratome or microtome, depending of what kind of microscopy you want to use later. Because the bacteria are tagged with fluorescent proteins, the best choice should be confocal laser microscopy.
If the bacteria are outside the tissues, on the surface, then you can't fix or treat the material in any way, because you will wash the bacteria from the surface. The best option should be confocal laser microscopy observations using fresh material and cut by hand.
I hope this can be helpful.
Thanks for your answers. In fact my goal to investigate their interaction not just in roots but in different parts of rice physiology (roots, leaves and stems). These bacteria are plant pathogen and are supposed to be inside the plant tissue. I'm searching all the protocols I'll need while the lab is getting the fluorescent vector kits, filters and other related materials for start this work. Your answer was very helpful.
It is very general question , first you have you to dehydrate the tissue (leaves, stem and root) in different grade of ethanol then this dehydrated tissue should be fixed in embedding medium (Sigma) and should be cut by using microtome of desired thickness.
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