uM molecule versus basepair calculation - (Dec/14/2010 )
Not really a homework question but not sure which forum I would ask calculation questions. I've been doing a bunch of calculations converting nanodrop ng/ul to uM and ran into an interesting dilemma.
Scenerio 1) Whenever purchasing primers, they usually come lyophilized in nmol that is calculated from A260 and converted with the epsilon value to get a nmol in molecule form, not nmoles of basepair so length matters. ie: 107 nmols = 107 nmols of molecules, not 107 nmols of bases.
Scenerio 2) When i convert dsDNA from nanodrop ng/ul, i usually convert by 660 g / 1 mol bp without taking into account the dsDNA length, giving a uM in bp rather than molecule. Units can still come out to uM meaning micromoles of basepair / L.
My question: Is it general practice to assume moles in DNA conversions automatically refer to moles of molecules rather than moles of basepair, and as such I should add in the length of the DNA in the calculation.
Example online calculators such as promega and ABI / roche take into account the length or molecular weight (which indirectly takes into account the length) and label the answer as pM for example, but don't specify pmoles of molecules / L or pmoles of basepairs / L. The calculations do correspond to pmoles of molecules / L but I wanted to see whether this is general practice throughout the community. Nanodrops give a tangible ng/ul value taking into account the physical bases via beers law does it not?
Thanks and hope it's not too confusing.
statements of concentration are always referring to molecules, not base pairs. if they want to speak about base pairs they will specifically say so.
Thanks for the reply. Is that just general accepted knowledge or is there a reference..or is that working off of chemistry where concentration is per molecule, not each carbon within the molecule kind of analogy?...just too used to being a scientist that's always asked to show "proof"....but thanks for the reply though!
Concentration is ALWAYS for the molecule unless specified, this is convention that has been around as long as we have been able to work out concentration and is regulated/standardised by IUPAC.
Unfortunatly you have been calculating your concentrations incorrectly - though they are easily converted if you know the length of your DNA/RNA.