Triturating plant tissue before isolation

Use this category for the exchange of ideas, methodologies and references regarding the isolation, manipulation and analysis of RNA. (Extraction protocols, Northern Blot analysis, RNase Protection, Differential Display, In Vitro Transcription, etc.)

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Triturating plant tissue before isolation

Postby wnt » Mar 14 2017 8:02 pm

Hello,

It's my first time in this place, because I've just started my adventure with molecular aspects of biology. So please, be tolerant :)

In last time I isolated plant tissue (roots, leaves), however the amount, which I got, was very low. Before isolation, I triturated plants' organs in liquid nitrogen. After that I did the isolation with Tri-reagent (Sigma) according the protocol. First time, when I got small account of RNA (200-300 [ng/┬Ál]), I changed reagents (I used new isoprophanol, ethanol, DEPC), but the problem didn't disappear. So, is it possible, that too many/too little amount of the liquid nitrogen, which I used during triturating organs, is the reason of my problem?

Other hand - I had to use organs from Cucumis sativus (Krak) - plants grew three weeks on full-medium, however during trird week on the leaves appeared chlorosis and nerosis. If it may be a reason of so low account of RNA from roots and leaves, which I isolated?

I will be very thankful for answers and suggests!
wnt
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Re: Triturating plant tissue before isolation

Postby nekameneka » Jun 16 2017 12:50 pm

Hi wnt,

Sorry for late reply, I hope you still read it. I would like to know why you use nitrogen to do that? I think you lost a lot of RNA in this process.
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