Hi everyone, i am doing mycoplasma detection by doing Pcr and im doing PCR 10ul reaction, my primer concentration os 2pmol and vol is 0.1ul. As i dont have pipette for 0.1ul i am thinking of diluting primers with water and taking highest concentration (dilution) . i dont quite understand this process. How much volume should i addand how much to take from the diluted vial so that i'll have concentration similar to 2pmol. Thank you in advance. Looking for your responses.
Try to make a master-mix (MM) if the number of samples is less and store the remaining, so that it can be used for the next set.
If the number of reaction are more you can always make one (MM).
Good Luck.
- Take 1ul of primer. Dilute with 1ul of water = 1 in 10 dilution then use 1ul of 1/10 dilution = 0.1ul of original
- However, let me stress your primer concentrations for standard PCR are WAY TOO LOW
- If it were me (having performed PCR since 1992) I would use 1ul neat of your primer @ 2uM = 2pmoles per ul
- As a minimum you are using about 10% of what you actually need for efficient PCR
C1V1=C2V2
Plug in the desired final concentration, final volume, and stock concentration. Solve for the volume of stock solution.
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