The answers in bold are correct, all I want to know is how to get those answers

Question

The answers in bold are correct, all I want to know is how to get those answers and the math involved depending on stock and fnial concentration.

15. To set up PCR last week, you used a solution containing everything except water, primers and template DNA. So, here you will work with some of the calculations required for putting together a PCR from "scratch". (3 pts) Stock 10X PCR Buffer 10 mM dNTPs (2.5 mM each) 25 mM MgClz 10 ?M Primer (each) 100 ?g/ml Template DNA 5 Units/ ?| Taq Polymerase sterile ddH20 (to make up volume How much of each of the following do you need to make 100 ?1 of Master Mix: Final Concentration 1X PCR Buffer 200 ?? dNTPs 1.5 mM MgCl2 0.1 uM Primers 200 ng Template DNA 1.25 Units of Taq 10 X PCR Buffer dNTP Stock Soln MgCl: Stock Soln Primers ul ul ul 2 6 or Taq Polymerase Sterile ddH2O Total Volume -56 or 55 ?! 100 ?

Explanation / Answer

As the question given that final reaction volume is 100ul. Hence you have to calculate the each and everything according to 100 ul volume.

Here stock of buffer is 10X mean it is 10 fold concentrated and you need 1X final concentration in 100 ul

Now calculate how much amount is of stock is required to make 1X solution of buffer for 100ul reaction.

You can you sinple

C1, V1 =C2 V2 formula

10 X * V1 ( volume is required)= 1X * 100

V1= 100/10= 10 ul

C1 V1= C2 V2

10 mM * V1= 200 uM * 100 ul

200uM= 0.2 mM

V1= 0.2 * 100/10

V1= 2 ul

Stock solution is 25 mM

25 mM * V1= 1.5 mM * 100

V1= 1.5 * 100/25

V1= 1.5 * 4= 6 ul

Stock solution10 uM or each primer ( forward and reverse)

10 uM * V1=0.1 * 100

V1= 10/10= 1ul

Here 1 ul of forward primer and 1 ul reverse primer.

Stock concentration

5 U/ul

5 U * V1= 1.25 * 100

V1= 1.25 * 100/5

V1= 125/5

V1= 25 ul

Here final volume is 100 ul so how much amount of water is required to make 100 ul final volume

100- ( 10 +2+6+2+25)

100-45= 55 ul

Here they if each primer is 0.5 ul is added then water volume will be56 ul.

If you have any doubt please ask. Kindly rate.

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