claritylight, on 27 January 2011 - 06:45 PM, said:
Sorry if I have been so unclear. I'm just saying there is 10ug of DNA at concentration 200ng/ul, so yes there's 50ul volume. So 1ug or 40ng/ul of DNA is in the final pcr reaction. The person I was talking to had got me all confused so I wanted to make sure I was right first of all. They were saying something about the total amount of DNA not changing in the reaction.
My other question is why is this way wrong: (5ul/25ul)*10ug = 2ug. Since the ul cancel out, you are left with ug only. Is this used for anything?
I know it's correct if you did it using ng/ul: (5ul/25ul)*200ng/ul = 40ng/ul.
You first need to clear about terms used.
amount = ug or ng ... so on
volume = ul or ml ... so on
Concentration = ug/ul or ng/ul or ug/ml ... so on.
The term "mass
" might be equal to amount coz mass refers to weight. But I think people don't use it often coz it's confusing.
The person you referred to; he is right. Total amount(ug or ng) of DNA in reaction doesn't change irrespective of PCR reaction volume. Only final concentration (ug/ul or ug/ul) changes.
If you add 5ul of 0.2ug/ul, there will be 1ug (1000ng) of DNA in your PCR reaction. That's what you called final amount.
If want to talk about final concentration, Yes, it does change.
If you PCR reaction volume is 20ul, the concentration will be 0.05ug/ul or 50ng/ul
If you PCR reaction volume is 50ul, the concentration will be 0.02ug/ul or 20ng/ul
Hope this helps.
P.S. Before you go on discussion, you guys should make it clear whether you all are on same page.
If you were talking about amount and others were talking about concentration, it'd be pointless arguing.