4 replies to this topic

[Marvin]

Hi,

I'm using a protocol that extract DNA from wood samples. For extraction it uses chlorofrom-isoamyl alcohol and phenol-chloroform-isoamyl alcohol and centrifuge at 10,000g for 10min for each extraction. However, we are using 50ml tubes for extraction so the table centrifuge doesn't fit. The maximum we can get with our large centrifuge is around 1,700g.

What I wonder is the function of the centrifuge, is 10,000g centrifuge just to make sure all wood residue be pushed down to the bottom or does it relate with the extraction rate?

Thanks.

[mdfenko]

it is to ensure that you pellet all of the wood and separate the aqueous from the organic layer.

[Marvin]

Thanks, mdfenko

I decide to try the slower centrifuge 1700g. But how can I know that the DNA is in the aqueous phase?

[Ameya P]

Marvin,

In the next step, where you transfer the supernatant and add iso-amyl alcohol, you should be able to see white strands if you mix the two liquid layers. That is an indication that you have some nucleic acids in the supernatant.

[mdfenko]

Marvin, on 05 April 2011 - 05:00 PM, said:

Thanks, mdfenko

I decide to try the slower centrifuge 1700g. But how can I know that the DNA is in the aqueous phase?

the dna enters the aqueous phase when you mix. the centrifugation is to speed up the separation of phases as well as to pellet solids.