Hi Eugene

Im not completely sure about this one - others may have definitive comments - but I have personally never precipitated DNA at such low speeds. I have had occasion to increase the centrifugation time from 15 min to 30 minutes and that might help; as will cooling the rotor to 4C

But in all honesty I would be wary and have never spun DNA at such low speeds. Therefore I will say no more but would be interested myself to hear from people who have had personal experience of low speed centrifugation in order to precipitate DNA

Hi Laurence

Thank you for your answer.  I think this is also something to consider.  Alternatively, based on the protocol, I am not sure if it is advisable to divide into 1.5-ml micro tubes of equal volumes after step 4 of "Isolation of DNA" and then proceed with the rest of the steps by adding an appropriate volume of the kit reagents. I say this because I have smaller centrifuge machines that are able to reach speeds of 8000 x g and higher.  However, my main worry for this is whether it is a totally wrong approach by correspondingly dividing the stated volume of the reagents in the protocol for the subsequent steps.  

Nevertheless, I would be interested in your thoughts for this.

Since you have a high expected quantity of DNA in your samples (at least 10's of µgs), splitting the volumes up into several tubes should not be an issue. Keep the high speed to get a quantitative precipitation, that is the preference here.

Only if you have starting material that will yield low DNA quantities will you need to be careful not to split the sample, as you need minimum quantities to get a precipitate to form during centrifugation. At very low quantities (ngs per tube), some people add e.g. commercial tRNA as a carrier for precipitation. Also, with very low quantities, sample loss by binding to the tube wall is an issue: then use special DNA low-binding tubes.

Hello again Eugene

I have just taken a look at the actual protocol and in essence I agree with Sven:

If this were me I would:

1. Add 3 volumes of cold ethanol to your lysate lysate

2. Incubate @ -20C for 15 min

3. Divide amongst a number of 1.5ml eppendorfs and then spin for 15 minutes at 13K in the microfuge.

This is what I routinely do and will result in good precipitation of your DNA

4. If necessary you can resuspend the pellets in TE or molecular grade (milli Q) water and then pool them back together having spun as aliquots in eppendorfs in a Microfuge

Hope that helps !! 

Noted!  Thank you Laurence and Sven for your contributions!

i have a c23 model cooling centrifuge which can reach only upto 8000 RPM which method to use for dna isolation from whole blood.