How to figure out the copy # per volume for a nucleic acid:
Essential information:
1. 1 ug of 1,000bp DNA = 1.52 pmole
2. Avogadro’s # = 6.022x10^23 molecules in a mole
3. Average molecular weight of a nucleotide is 327.0
4. Have to know the size of the nucleic acid (and if it is double or single stranded)
Example question: Let’s say you have a solution of 498ng/ul plasmid DNA and that the plasmid is
3,260bp in size. You want to determine how many copies of the plasmid there are per microliter so that you can make a Q-PCR standard curve of this plasmid.
1. What is mass of a single nucleotide?
Average molecular weight of a single nt = 327 (g/mole)
A single nt is only 1/6.022x10^23 of a mole
(327g/mole)/(6.022x10^23molecules/mole)=5.43x10^-22 g/molecule
2. So, what is the mass of a single copy of the plasmid?
However, you have 6,520 nucleotides in a 3,260bp plasmid
5.43x10^-22 g/molecule x 6,520 molecules = 3.54x10^-18g = mass of a single plasmid copy
This is the equivalent of 3.54x10^-9ng per single plasmid copy
3. If you have the concentration of your plasmid stock, use that to figure out the plasmid cop #:
Since your solution is 498 ng/ul, here is how you calculate the number of plasmid copies per ul:
1 plasmid copy / 3.54x10^-9 ng x 498ng/1ul = 1.41x10^11 copies per ul