DNA Ligation and Bacterial Transformation
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DNA Ligation and Bacterial Transformation |
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| Subject
Area |
AP Biology |
| Age
or Grade |
11-12 |
| Estimated
Length |
The entire experiment takes four days, however not all days require the entire class period. |
| Prerequisite
knowledge/skills |
Students should have previous knowledge of the structure of a plasmid, the function of retriction endonucleases, the general process of DNA ligation ("sticky ends"), as well as the concept of bacterial transformation. |
| Description of New
Content |
Students will learn basic molecular techniques in forming recombinant DNA using precut plasmids, as well as plating and growing transformed bacterial cells. Students will be introduced to the concepts of selection using antibiotic resistance, as well as the lac operon and the function of the β-galactosidase gene. |
| Goals |
To help students better understand the concepts of DNA ligation and transformation using a hands on approach in which they perform basic techniques in molecular biology. |
| Materials
Needed |
Science Vision International cloning and transformation kit, incubator, orbital shaker, micropipettes, hot plate, autoclave (ideal, but not necessary), various glassware (beakers, etc), innoculating loop/flame. |
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Procedure
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Opener: Students should be given a pre-lab introduction in which they read and outline the rationale for the experiment. Development: Students should conduct the ligation and transformation experiments in small groups (2-3) by following the instructions given by the teacher. Students should keep a detailed log of procedures/methods as they work each day. There will be a minimal amount of teacher prep time before each lab period. Closure: Students should record the number and color of colonies formed on each of their plates. Questions are included in the instruction handout above. Compilation of data followed by class discussion/analysis of these results is helpful as well. |
| Evaluation |
Students should hand in their log kept during the course of the experiment as well as their detailed results and the answers to any questions included in the instruction packet. It is important that they are able to list possible sources of experimental error (not including human error). |
| Extensions |
An additional lab could be added in which students run the DNA fragments (pre-cut with restriction endonucleases, comes in the kit) through gel electrophoresis to visualize the size of fragments digested with different restriction enzymes. |
| References | Science Vision International cloning and transformation kit |