BISC 219/2009:Creating a Transgenic Organism: Difference between revisions
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##measuring the activity of the introduced enzyme, β–glucuronidase (GUS) spectrophotometrically and histochemically | ##measuring the activity of the introduced enzyme, β–glucuronidase (GUS) spectrophotometrically and histochemically | ||
##PCR to test directly for the presence of the introduced genes and <br> | ##PCR to test directly for the presence of the introduced genes and <br> | ||
#become adept at sterile technique. | #become adept at sterile technique. | ||
{| border="1" | |||
|+ | |||
! Concepts !! Genes/Organisms !! Techniques/Skills | |||
|- | |||
| Effect of random gene insertion in a large genome | |||
| Bacterial pathogen: ''Agrobacterium tumefaciens'' || Aseptic tissue culture | |||
|- | |||
| Plant totipotency | |||
|Tobacco plant: ''Nicotiana tobacum'' | |||
|DNA Extraction | |||
|- | |||
|Regulation of gene expression | |||
|gusA gene from ''E. coli'' | |||
|PCR | |||
|- | |||
|Genetic Engineering and creation of genetically modified organism | |||
| | |||
|Agarose gel electrophoresis | |||
|- | |||
|Phenotypic Selection | |||
| | |||
|Transfection & selection by antibiotic resistance; | |||
Enzyme function assays: colorimetric & histochemical | |||
|} |
Revision as of 12:44, 13 April 2009
At the end of this semester long series you will have mastered the following skills and concepts using both microorganisms and a multicellular organism.
Schedule of Experiments
Lab # | Dates | Activity |
---|---|---|
1 | 9/2 - 9/8 | set-up of plant transformation |
Independently | 9/5 - 9/13 | transfer of explants to selective medium; induce shoots |
5 | 9/30 - 10/6 | transfer shoots to root inducing medium |
7 | 10/15 - 10/21 | transfer plantlets to soil |
9 | 11/12 - 11/18 | GUS activity assays; set up PCR reactions |
10 | 11/19 - 11/25 | Gel electrophoresis of PCR product |
11 | 12/9 | Plant Lab Report Due for all students |
Experimental Objectives:
In this semester long experiment you will:
- use the Agrobacterium system to introduce a foreign gene, the β -glucuronidase gene (gusA) of E. coli, into the cells of tobacco
- take advantage of the totipotency of plant cells and use these transformed cells to regenerate genetically engineered plants
- confirm that the regenerated plants are transformed by
- measuring the activity of the introduced enzyme, β–glucuronidase (GUS) spectrophotometrically and histochemically
- PCR to test directly for the presence of the introduced genes and
- become adept at sterile technique.
Concepts | Genes/Organisms | Techniques/Skills |
---|---|---|
Effect of random gene insertion in a large genome | Bacterial pathogen: Agrobacterium tumefaciens | Aseptic tissue culture |
Plant totipotency | Tobacco plant: Nicotiana tobacum | DNA Extraction |
Regulation of gene expression | gusA gene from E. coli | PCR |
Genetic Engineering and creation of genetically modified organism | Agarose gel electrophoresis | |
Phenotypic Selection | Transfection & selection by antibiotic resistance;
Enzyme function assays: colorimetric & histochemical |