953 - EDVOTEK

22
EDVO-Kit #
953
Water Quality Testing III: Multiplex PCR Analysis for Water Contaminants
Pre-Lab Preparations
Notes and Reminders:
Instructor’s Guide
• Accurate temperatures and cycle times are critical for PCR. A pre-run for
one cycle (approx. 3 to 5 minutes) is recommended to check that the thermal
cycler is properly programmed.
• For thermal cyclers that do not have a top heating plate, it is necessary to
place a layer of wax above the PCR reactions in the microcentrifuge tubes to
prevent evaporation. See Appendix entitled "Preparation and Handling PCR
Samples with Wax ".
• Three water baths can be used for PCR if a thermal cycler is unavailable.
The experiment will require great care and patience. Samples will require
wax layers. See appendices entitled "Polymerase Chain Reaction Using Three
Water baths" and "Handling samples with wax overlays".
MODULE II - AMPLIFICATION OF THE Multiplex Water
Contaminants
The Thermal cycler should be programmed for the following cycles:
Initial Denaturation 35 cycles @ Final Extension
94°C for 5 min. 94°C for 30 sec. 72°C for 4 min.
58°C for 30 sec.
72°C for 60 sec.
• Accurate temperatures and cycle times are critical. A pre-run for one
cycle (which will take approximately 3 to 5 min) is recommended to
check that the thermal cycler is properly programmed.
• For thermal cyclers which do not have a top heating plate, it is necessary
to place a layer of wax above the PCR reactions in the microcentrifuge
tubes to prevent evaporation.
• Three water baths can be used for PCR if a thermal cycler is unavailable.
The experiment will work if given great care and patience. Samples will
require wax layers.
Duplication of this document, in conjunction with use of accompanying reagents, is permitted for classroom/laboratory
use only. This document, or any part, may not be reproduced or distributed for any other purpose without the
written consent of EDVOTEK, Inc. Copyright © 2006, 2008, 2009 EDVOTEK, Inc., all rights reserved
EVT 090205K
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