Chapter 3 Prepare the Reactions
Prepare the Standard Dilution Series
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Standard Curve
Experiments
48
Notes
2. Add the required volume of water (diluent) to each empty tube:
3. Prepare dilution 1 in the RNase P Std. 1 tube:
a. Vortex the stock for 3 to 5 seconds, then centrifuge the tube briefly.
b. Using a new pipette tip, add 9.08 µL of stock to the RNase P Std. 1 tube.
c. Vortex Std. 1 for 3 to 5 seconds, then centrifuge the tube briefly.
4. Prepare dilution 2 in the RNase P Std. 2 tube:
a. Using a new pipette tip, add 9.08 µL of dilution 1 to the RNase P Std. 2 tube.
b. Vortex Std. 2 for 3 to 5 seconds, then centrifuge the tube briefly.
5. Prepare dilution 3 in the RNase P Std. 3 tube:
a. Using a new pipette tip, add 9.08 µL of dilution 2 to the RNase P Std. 3 tube.
b. Vortex Std. 3 for 3 to 5 seconds, then centrifuge the tube briefly.
6. Prepare dilution 4 in the RNase P Std. 4 tube:
a. Using a new pipette tip, add 9.08 µL of dilution 3 to the RNase P Std. 4 tube.
b. Vortex Std. 4 for 3 to 5 seconds, then centrifuge the tube briefly.
7. Prepare dilution 5 in the RNase P Std. 5 tube:
a. Using a new pipette tip, add 9.08 µL of dilution 4 to the RNase P Std. 5 tube.
b. Vortex Std. 5 for 3 to 5 seconds, then centrifuge the tube briefly.
8. Place the standards on ice until you prepare the reaction plate.
Preparation
Guidelines
When you prepare your own standard curve experiment:
• Standards are critical for accurate analysis of run data.
• Any mistakes or inaccuracies in making the dilutions directly affect the quality of
results.
• The quality of pipettors and tips and the care used in measuring and mixing
dilutions affect accuracy.
• Use TE buffer or water to dilute the standards.
Tube Standard Name Volume of Diluent to Add (µL)
1 RNase P Std. 1 9.08
2 RNase P Std. 2 9.08
3 RNase P Std. 3 9.08
4 RNase P Std. 4 9.08
5 RNase P Std. 5 9.08
To Next Page
Loading...
Need help?
General
