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Give your students the opportunity to learn and explore transcription regulation right in your classroom. This unique approach to synthetic biology was developed by college professors focused on creating a unique activity to demonstrate gene regulation. This multi-part lab will expose students to cloning, restriction enzymes, transformation, microbiology, and so much more in an effective classroom protocol.
AdvancedFor experienced high school and college classes; requires some technical skill.
Bring the unique area of synthetic biology into your classroom with this one-of-a-kind kit activity. Your students get the opportunity to take on the role of genetic engineers as they clone promoters into 2 different plasmids. This hands-on approach to transcription regulation exposes your class to a wide array of biological disciplines including biotechnology, molecular biology, and evolutionary biology.
Students clone the promoters into the plasmids, pClone Red and pClone Blue, using a common cloning technique, Golden Gate Assembly (GGA). After cloning in the promoters, students are tasked with performing a transformation on E. coli cells to produce new ampicillin-resistant colonies expressing their designed plasmids. Based on the orientation of the promoter, students then observe colonies that appear green (produce GFP), red (produce RFP), or blue (produce AmilCP blue). This gives them 3 distinct colonies to further assess using ImageJ software, freely available, to determine the level of protein expression.
The kit includes enough materials for 8 groups of 2 to 4 students. The activities typically take 3 to 6 class periods for completion. Note: Kit comes with the perishable materials included.
|Ampicillin, 10 mg/mL||4||Included|
|E. coli with Plasmid Containing Plac Promoter and RFP Gene||1||Included|
|E. coli with Plasmid Containing P5 Promoter and AmilCP Blue Gene||1||Included|
|T4 DNA Ligase||1||Included|
|10x Ligase Buffer||1||Included|
|MM294 E. coli Slant||1||Included|
|Mutant Oligonucleotide Pair (P5-33A top and bottom)||1||Included|
|Wild-Type Oligonucleotide Pair (P5 top and bottom)||1||Included|
|pClone Red Plasmid, 25 µg/µL||1||Included|
|pClone Blue Plasmid, 25 µg/µL||1||Included|
|10x Oligonucleotide Annealing Buffer||1||Included|
|Calcium Chloride, 50 mM, 3 mL||8||Included|
|Disposable Spreaders, Sterile||30||Included|
|Disposable Inoculating Loops, Sterile||30||Included|
|LB Agar, Sterile, 400 mL||4||Included|
|LB Media, 3 mL||8||Included|
|Transformation Tubes, 15 mL||25||Included|
|Digital Resource Instruction Card||1||Included|
|Teacher’s Manual with Reproducible Student Guide||1||Included|
|Aluminum Foil||Needed, Not Included|
|Beaker, 1 L||Needed, Not Included|
|Computer||Needed, Not Included|
|Digital Camera||Needed, Not Included|
|Disinfectant (ethanol or household bleach solution)||Needed, Not Included|
|Hot Plate or Bunsen Burner||Needed, Not Included|
|Ice||Needed, Not Included|
|ImageJ Software (free download)||Needed, Not Included|
|Incubator, 37° C (optional)||Needed, Not Included|
|Lab Marker||Needed, Not Included|
|Microcentrifuge||Needed, Not Included|
|Microcentrifuge Tubes, 0.5 mL||Needed, Not Included|
|Micropipettors||Needed, Not Included|
|Micropipette Tips, Sterile||Needed, Not Included|
|Parafilm®||Needed, Not Included|
|PCR Tubes, 0.5 or 2.0 mL||Needed, Not Included|
|Programmable Thermal Cycler||Needed, Not Included|
|Thermometer||Needed, Not Included|
|Tongs||Needed, Not Included|
|Tube Racks, for 0.5-mL Tubes||Needed, Not Included|
|Tube Racks, for 15-mL Tubes||Needed, Not Included|
|UV Transilluminator||Needed, Not Included|
|White-Light Transilluminator (optional)||Needed, Not Included|
|Water||Needed, Not Included|
|Water Bath, 42° C||Needed, Not Included|
|Wire Inoculating Loop||Needed, Not Included|