miniPCR bio
TM
Chopped! Using CRISPR/Cas9 to cut DNA - Instructor's and Student's Guide
Version: 1.0 - Release: May 2022 - © 2022 by miniPCR bio™
P./7
Instructor's Guide
Lab setup
The following activities should be carried out by the instructor ahead of class. Reagents are
sufficient for 8 student groups.
Preparation for optional pre-lab activity
• Print the pre-lab activity from the lab guide (pages 40-42). Students will be cutting out
materials from these pages, so be sure to print single-sided!
• Provide each student group with scissors.
• Note: The pre-lab activity in the guide instructs students to cut out paper models of the gRNA
and DNA to determine the expected band sizes. If you do not wish to use the paper model
cutouts, you can have your students analyze the sequences electronically with the provided full
sequences (available at under the downloads tab at
https://www.minipcr.com/product/chopped-crispr-cas9-lab/) or skip the pre-lab altogether
and just provide the expected band sizes to your students (found on page 44).
Preparation for in vitro CRISPR/Cas reaction
Important notes about reagent handling:
• The most common source of failure in this activity is due to degradation of the Cas9 Nuclease
due to incorrect handling. Keep the Cas9 Nuclease in the freezer or on crushed ice at all
times.
• 5X Cas9 Nuclease Concentrate will remain liquid at -20 °C and does not need to be thawed.
• The Cas9 nuclease is supplied as a 5X concentrate. Be sure to dilute the Cas9 enzyme as
described on the following page.
• All other reagents are stable at room temperature for at least one hour but should remain cold
or on ice for longer periods and frozen for long-term storage.
Gloves and protective eyewear should be worn for the entirety of this experiment.
Keep the Cas9 Nuclease on ice at all times.
