Knockout! A CRISPR/Cas Gene Targeting Lab
This lab is easy for teachers to prepare, makes gene targeting accessible to students, and packs a powerful punch!
Become a genetic engineer and use CRISPR/Cas to disable a gene in
bacteria. Scientists use the CRISPR/Cas system to alter the DNA in
living cells. Now you can bring this cutting-edge technique to your
a
dvanced biology class! In this lab, E. coli cells which express the lacZ gene from a plasmid will be transformed with another plasmid containing the cas9 gene and a custom guide RNA. This guide RNA contains a sequence complementary to the lacZ gene so, in transformed cells, the guide RNA will target Cas9 to cut the lacZ
gene, disabling it. Students can easily identify their CRISPR/Cas
knockouts using the color of the bacterial colonies. Cells which express
lacZ are able to break down the chemical X-gal into a blue product. Cells with a functional lacz will be blue, while cells in which the CRISPR/Cas system has successfully disabled lacZ will revert to a white phenotype.
Note: This is a technically advanced lab that
requires a 2-hour class period. Students should have prior experience
with bacterial transformation.
- Techniques: Micropipetting, bacterial transformation, gene targeting
- Topics: Genome editing, biotechnology, molecular biology
- Time required: Requires a 2-hour class period
- Level: Advanced high school through college
This lab provides enough reagents for eight lab groups:
Required equipment:
- Micropipettes and tips
- Water bath or dry bath
- Incubator
Storage notes:
- Some reagents require refrigerator storage and some require freezer storage
- Reagents are stable for 3 months when stored as directed
Refill kit available:
- Does not include necessary plasticware and LB-agar media,
Knockout! A CRISPR/Cas Gene Targeting Lab (Refill kit)
Refill kit, does not include necessary plasticware and LB-agar media.
This lab is easy for teachers to prepare, makes gene targeting accessible to students, and packs a powerful punch!
Become a genetic engineer and use CRISPR/Cas to disable a gene in
bacteria. Scientists use the CRISPR/Cas system to alter the DNA in
living cells. Now you can bring this cutting-edge technique to your
advanced biology class! In this lab, E. coli cells which express the lacZ gene from a plasmid will be transformed with another plasmid containing the cas9 gene and a custom guide RNA. This guide RNA contains a sequence complementary to the lacZ gene so, in transformed cells, the guide RNA will target Cas9 to cut the lacZ
gene, disabling it. Students can easily identify their CRISPR/Cas
knockouts using the color of the bacterial colonies. Cells which express
lacZ are able to break down the chemical X-gal into a blue product. Cells with a functional lacz will be blue, while cells in which the CRISPR/Cas system has successfully disabled lacZ will revert to a white phenotype.
Note: This is a technically advanced lab that
requires a 2-hour class period. Students should have prior experience
with bacterial transformation.
- Techniques: Micropipetting, bacterial transformation, gene targeting
- Topics: Genome editing, biotechnology, molecular biology
- Time required: Requires a 2-hour class period
- Level: Advanced high school through college
Lyophilized E. coli for Knockout! Lab( optional add on)
Optional add-on to carry out transformations on different days.
One vial of lyophilized E. coli cells, optional add-on to the Knockout! CRISPR/Cas Lab.
In the Knockout! Lab, bacteria are transformed with the pKO plasmid (encoding Cas9 and a guide RNA that targets the lacZ
gene) or with the pCtrl plasmid (encoding Cas9 and a random guide RNA).
Once reconstituted in LB-Amp liquid media, the lyophilized bacteria
must be transformed within 48-72 hours. Adding a second vial of
cells allows you to carry out transformations on different days with
different lab groups, thus making it feasible to split the kit into lab
groups working on different weeks.
Please refer to the Knockout! Lab Instructor’s Guide for detailed instructions.
This product is sold for educational use only. It contains technologies that are owned and licensed for educational purposes only by miniPCR bio.
This kit is an optional add-on to the Knockout! Lab
(MP-KT-1800-01 or MP-KT-1800-03).
One vial of cells allows you to split
the Knockout! lab into separate transformations at different times. One
vial contains enough cells for 8 lab groups.
Storage notes:
- Cells may ship at room temperature, but require fridge storage upon arrival
- Cells are stable for 6 months when stored in the fridge
| Class |
AP Biology, Biotechnology
|
|---|
| Grade level |
Advanced High School, College
|
| Technique |
PCR, CRISPR/Cas |
SDS Lyophilized DH5-alpha E. coli
If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200