Hi, Annalisa,

generally, one can got up to 0,2 µg of the DNA pre mg of the tissue. If you have use old leaves with large cells, the amount are less.

I guess you will need at least 100 g of the tissue to get 1mg.

Goog luck!

Hi! Yes the leaves were quite old..I used 400 milligram and got moreless 150 micrograms then yes I think I have to use at least 100-200 grams of this material...

young materials are necessary. fine ground (so much leaves, you can use a machine), add more water into the tube after chloroform extraction and re-do chloroform extraction (just insure not much DNA lost).

if you take old leaves, you need to crush it more and have to take more leaf sample as said above ,repeat the addition of Chloroform : Iso Amyl Alcohol (24:1) once more, sum up the upper aqueous phase and add Ammonium Acetate, ethanol accordingly. After adding ethanol invert tubes very gently so that the total precipitate will be altogether.

thank you all. yes in fact I have tried with more material (800 mg of crushed Leaves) and I am doing the chloroform extraction twice. so I get a cleaner , but however dark green DNA pellet. The amount until now is moreless 2-3 ml of 200 ng/ul of DNA solution...not so bad but with a quite low A260/A280 ratio....1.3-1.4

CTAB method

Follow the method on Plant Methods 8:26. I routinely used freeze-dried leaf samples from 20 to over 100 mg. I obtain 50 to 200 micro gram of DNA. You can use large preps to obtain mg of pure DNA

CTAB is fine. We can have high concentration DNA for whole genome sequencing.

I agree with CTAB method.

Mace et al., 2006 has an excellent DNA extraction protocol using CTAB

Hi Annalisa,

To get large amount of DNA from plant tissue using CTAB method you should take care, that plant tissue should be young and grounded properly and to powdered tissue add CTAB buffer kept at 65 degree, and tissue should be mixed occasional swirling of tubes so no tissue remain attached to tube.In chloroform:Isoamyl step two layers should be mixed properly and and on addition of isopropanol tubes should be gently swirled so DNA threads remain intact.

Detailed information about DNA quantity and tissue amount is given in Doyle and Doyle 1990 paper in Focus volume 12, page number 13-15. this paper's discussion is really valuable