A master mix (sometimes called a cocktail) is a single tube that contains all of
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A master mix (sometimes called a cocktail) is a single tube that contains all of the components that are common to multiple tubes. The purpose of the master mix is to minimize the number of solutions that you need to pipette and to minimize the possibility of pipetting errors. It also helps avoid having to pipette very small volumes Fill out the following chart to determine the volumes of each solution required for your master mix The final PCR reaction volume is 25 The first column described the final concentrations of each solution in the master mix The second column is the stock, or the concentration of the tubes you are given to make the master mix The third column is the volume required for one reaction (xxn), or a single tube containing the master mix plus one DNA sample you are testing The fourth column represents the volume required to make ten reactions, each with different DNA samples Show your work below the chart. (10 points) 10 Yol Stock Yol 10X 1X PCR reaction buffer (contains 1.5 MMgCl.) 200 uM solution with all four dNTPs (pH 8.0) 0.8 HM, forward primer 0.8 uM reverse primer 0.05 units (U) thermostable DNA polymerase dH2O 12.5 uM 12.5 uM Show vour work below.Explanation / Answer
stock 1rxn vol
1X PCR reaction buffer 10X 2.5 l.( 10/1=10 fold diuted; 25/10l)
200M DNTP mix 10mM 0.5 l ( 10 X 1000 M/ 200 M= 50 fold; 25/ 50l)
0.8 M forward primer 12.5M 1.6l ( 12.5/0.8=15.6 fold; 25/ 15.6 l=12.5l)
0.8M reverese primer 12.5M 1.6l ( 12.5/0.8=15.6 fold; 25/ 15.6 l= 12.5l)
0.05 U DNA polymerase 5U 0.25 l( 5/0.5= 100; 25/ 100 l= 0.25l)
dH20 18.55l
for 10rxn volume
1X PCR reaction buffer 25 l.
200M DNTP mix 5 l
0.8 M forward primer 16l
0.8M reverese primer 16l
0.05 U DNA polymerase 2.5 l
dH20 185.5l
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