Question

You have a sample of ssDNA molecule that you will use as a primer for PCR. You dissolved the dried ssDNA peller in TE to a total volume of 400 ul to make a "stock DNA sample." You need to determine the concentration (ug/ul) of this primer DNA in solution. To do this you took 4 ul from the "stock DNA sample" and added these to 996 ul of TE buffer in a cuvette, making a total of 1000 ul in a cuvette. Next, you determined the absorbance of 260 nm light for this 1 ml of diluted DNA in the cuvette and the A260 value is 0.014.

1) what is the concentration (ug/ul) of the DNA solution in the cuvette?

2) what is the concentration (ug/ul) of the primer DNA in the original Stock DNA sample?

3) what is the total amount (ug) of DNA in the stock DNA sample tube?

4) how many ul of the original "stock DNA sample" would you need to use in a PCR experiment that required a total of 0.25 ug of DNA from this sample?

Answer 1

Considering 1 OD = 1 g/L

1. Concentration of the DNA solution in the cuvette = 0.014 g/L

2. Dilution = 4/1000 = 0.004

Therefore, Concentration of the primer DNA in the original Stock DNA sample = 0.014/0.004 = 3.5 g/L

3. Volume in the stock tube = 400 L

So, total amount (g) of DNA in the stock DNA sample tube = 3.5 g/L x 400 L = 1400 g

4. Amount of original stock required for a total of 0.25 g of DNA from this sample = 1/3.5 x 0.25 = 0.0714 L