I've used Qiagen kits to isolate RNA from trees (leaves) and found them to be very effective and simple to use. They are column/membrane based and I managed to get excellent quality RNA for RNA-seq. I have heard that AMBION is also good for but as I got what I needed from Qiagen I never had to use another kit.

My laboratory has been using miRVana for different plants. It works very well.

I can recommend SPLIT kit from Lexogen http://www.lexogen.com/products/split-rna-extraction/split-description.html . It came out on the market recently, so maybe not that known yet. It works pretty good for plant RNA. This kit was optimized to get high quality RNA, because our downstream application is RNAseq. 

You can extract total RNA or small (cut off is around 150 nt) and large fractions. There's no DNAse treatment, but it efficiently gets rid of gDNA. It takes about 30 min for extraction. And it's not expensive kit.

I also recommend MirVana kits. However, I do find that I have to use 15ml tubes in part of the protocol, which is annoying. 

Our lab uses a protocol adapted from White and Kaper (1989).I find the yield to be much higher and is often quicker than the MirVana kit. see attached

I just want to isolate total RNA once and not repeatedly for different procedures like small RNA enrichment. This is to avoid any variations between samples, so the same RNA will be used for RNA-seq and small RNA-seq. Any kind of enrichment of one over the other may alter our data to some extent. So I guess these kits help isolate all kinds of RNA molecules including both long (mRNA) and small RNAs, without any biased enrichment of one over the other, unless we choose to do that way? 

WIth the SPLIT kit that's correct. When you extract total RNA, there's no preference over certain length. Well, technically there's always some variation and loss, but when a spiked-in RNA marker of length 200-10000 nt was mock-extracted, there was no obvious bias towards size. There's more info here http://www.lexogen.com/fileadmin/uploads/pdfs/SPLIT/008.048_SPLIT_AN_008AN019V0100_2014-08-27.pdf

I would suggest you use Tri-reagent if you can. After RNA precipitation step and getting RNA pellet, you can use Licl(4M) to wash the pellet in order to isolate small molecular RNAs including siRNA and miRNA. The pellet leftover will be large molecular RNAs. Precipitate the smRNAs with iso-propanol, wash with 80% ethanol and solve it into DEPC water. There is no limitation for starting materials and you can easily see the pellet after precipitation of smRNAs if you start with 10 g of tissue.

i've been using RNAeasy mini kit to isolate total RNAs and miRvana kit to isolate miRNA ! worked well, but i had to improve the miRvana protocol (made my own one) to improve the efficiency of the extraction !

Good luck :)

Qiagen kits to isolate Total RNA has proved to be the best for me

EIther QIagen's miRNeasy kit or Zymo's Direct-zol RNA purification kit will isolate both large and small RNA without a separate small RNA enrichment step.  Smart move avoiding small RNA enrichment, typically that is not advised because it is hard to accurately quantitate the concentration of a small RNA fraction.  Either yield high quality RNA although Zymo's kit eliminates the organic phase separation step.

I have used both kits and been happy with the results from both.  Zymo's kit is much cheaper than Qiagen's kit if cost is an issue.

 The only plant specific part you need to worry about is the homogenization step. How do you plan to homogenize the seedlings?  Possibly, bead-based homogenization using Zirconia beads (~2.8 mm)?

FYI - Zymo may send you a free sample of their Directzol kit.  Check their website.

Hi, Thanks for your answers. Many people have recommended using Qiagen's miRNeasy kit. But their handbook specifically says to use for purification of total RNA, including miRNA, from animal and human cells and tissues. Unlike their other versions (like RNeasy Plant Mini Kit) which are plant specific, this product handbook doesn't say like this and only mentions to use for animal and human cells, so how do we use this kit for plants?

I have not worked with plant tissue in a long time but I remember disruption and homogenization being at lot more complicated due to the presence of cell walls.  The major difference between the plant specific protocols and animal cell protocols are the disruption and homogenization methods (assuming your sample is compatible with guanidine thiocyanate).  For plants Qiagen recommends disruption with a liquid nitrogen cooled mortar/pestle, placed in lysis buffer (for miReasy this is phenol/guanidine) then homogenization with a Qiashredder (rotor-stator).  The plant kits include the Qiashredder but Qiagen mentions that bead-based homogenization techniques work as well.  Adapt the disruption, lysis, and homogenization techniques to your sample and after this simply follow the miRNeasy protocol and everything should work fine.

Hi Brian, 

I tried Qiagen miRNeasy kit for plants. The buffers and the RNA purification column for this kit is exactly the same as the RNeasy Plant Mini Kit. The plant kit uses different isolation buffer but the miRNeasy kit uses traditional trizol/chloroform method. They provide Qiazol which is similar to trizol. I talked to Qiagen, they said they haven't tried their miRNeasy kit for plants but I can test. I tested, I crushed the tissues in liquid nitrogen with mortar and pestle and centrifuged the plant debris in trizol (instead of using Qiashredder). I took the supernatant and then added chloroform and then following the protocol, added the aqueous phase to the column. The RNA readings, both 260/280 and 260/230 ratio came out well (above 2). Qiagen concern was the presence of plant polysaccharides and other compounds to be removed by miRNeasy kit but I think they came out well as the cleaning column and cleaning buffers are the same as the regular plant kit. So I think miRNeasy kit can be used and I hope they give good small RNA species along with the longer RNA fragments, isn't it? My other concern was if miRNeasy is isolating less amount of small RNAs, but I feel it should be OK. Since the input solution for both plant and animal is the aqueous phase of chloroform extraction, plant tissues should have the same amount of small RNAs as from the animal tissues. I believe and miRNeasy kit should be OK for plants, isn't it?

The kit works well for isolating large and small RNAs.  I have used it for assaying both miRNAs and mRNAs by qRT-PCR and I recall seeing a review article a while back that compared miRNeasy to RNeasy isolation and observed that miRNeasy worked best for isolating both small and large RNAs.  The RNeasy kit lost a good bit of the small RNA fraction while miRNeasy retained it while also isolating comparable amounts of the large RNA fraction.  Years ago I used phenol chloroform extractions for protein extractions from plant tissue with no issues but not RNA.  If you are concerned about polysaccharides, perhaps try running a denaturing agarose gel first to determine the purity of you isolates.  Perhaps additional washing or a second purification step will help remove polysaccharides.  Your absorbance at 230 will give you an indication of polysaccharide contamination so if your 260/230 ratio is decent perhaps polysaccharide contamination is not an issue in your case.  Either way, it always helps to visualize sample purifty using a denaturing agarose gel.  If this approach does not work it looks like Ambion has a Plant RNA isolation aid that is compatible with the miRVana miRNA isolation kit which supposedly isolates both large and small RNAs as well.