biomath question - please help a newbie - (Feb/19/2006 )

Hi,

Hi all,

I am doing a summer internship in a lab, and have got a problem that is quite embarrassing => my bad biomaths Could someone help me out with this seemingly simple calculation?

Stock solution Volume (ul)
dH2O 835
10 PCR buffer (inc. Mg2+) 100
dNTP mix (10 mM each) 20
Sense Primer 20
Antisense Primer 20
Taq pol (5U/ul) 5
-------------------------------------------------------
Total 1000 ul

Now, one will typically add 20 of this mix/well, however, my situation is different:

10 ul H2O already in the well, so need to make 2X premix and add 10 ul the 2x premix to the well in order to obtain the same conc. as if I added 20 ul of the 1000 ul premix above to a well. For some reason I cannot work this out..

Thankyou very much.

FZ

In your master mix, add half the water and leave everything else the same for a 2X solution.

Hi,

Hi all,

I am doing a summer internship in a lab, and have got a problem that is quite embarrassing => my bad biomaths Could someone help me out with this seemingly simple calculation?

Stock solution Volume (ul)
dH2O 835
10 PCR buffer (inc. Mg2+) 100
dNTP mix (10 mM each) 20
Sense Primer 20
Antisense Primer 20
Taq pol (5U/ul) 5
-------------------------------------------------------
Total 1000 ul

Now, one will typically add 20 of this mix/well, however, my situation is different:

10 ul H2O already in the well, so need to make 2X premix and add 10 ul the 2x premix to the well in order to obtain the same conc. as if I added 20 ul of the 1000 ul premix above to a well. For some reason I cannot work this out..

Thankyou very much.

FZ

thank you all

FZ

QUOTE (klugman @ Feb 20 2006, 07:22 AM)

Hi,

Hi all,

I am doing a summer internship in a lab, and have got a problem that is quite embarrassing => my bad biomaths Could someone help me out with this seemingly simple calculation?

Stock solution Volume (ul)
dH2O 835
10 PCR buffer (inc. Mg2+) 100
dNTP mix (10 mM each) 20
Sense Primer 20
Antisense Primer 20
Taq pol (5U/ul) 5
-------------------------------------------------------
Total 1000 ul

Now, one will typically add 20 of this mix/well, however, my situation is different:

10 ul H2O already in the well, so need to make 2X premix and add 10 ul the 2x premix to the well in order to obtain the same conc. as if I added 20 ul of the 1000 ul premix above to a well. For some reason I cannot work this out..

Thankyou very much.

FZ

For a two fold mastemix use the same stuff but at only 335µl H20. This will reduce the volume to 500µl so the concentration of everything contained in this mix will be doubled