On a separate sheet of paper, define the following terms. Terms are in the order they appear in the text.
nucleic acid probe
12.1 Summary: Transformation and Gene Transfers
- Genetically modified organisms are the product of intentional transformation, a process known as transgenics or gene transfer.
- DNA transformation begins by identifying the gene of interest and cutting it from its source through the use of restriction enzymes isolated from bacteria.
- Restriction enzymes recognize and cut specific DNA sequences at restriction sites, producing either blunt or sticky ends and small DNA pieces called restriction fragments.
- Once the gene of interest has been removed from the source DNA, a method of insertion into a target cell must be provided.
- Small circular DNA plasmids act as vectors. The plasmid is opened with the same restriction enzyme used to cut the source DNA, producing recombinant DNA.
- Target cells must be heat shocked or subjected to brief electrical shocks by electroporation to encourage the cells to take up the recombinant plasmids. Large eukaryotic cells can have the DNA injected into them using microscopic needles. Viruses may also be used as vectors to deliver genes.
- Once the target cells have taken up the gene of interest, they are encouraged to grow. This is the cloning of the cell and the introduced DNA.
- Not all target cells take up the vector with the gene of interest, so a screening process is used. Often, plasmids for antibiotic resistance (R) are used as the vector, so only cells with the R plasmid will grow on petri dishes containing the antibiotic.
- Some of the screened cells will possess plasmids without the gene of interest, so a second screening process must be used. Cells are transferred from petri dishes to filters to make a copy of the surface of the plate.
- The DNA of transferred cells is opened with chemicals or heat. A nucleic acid probe, complementary to the gene of interest, is added. It will be taken up only by cells with the gene of interest, allowing detection.
- Gene transfer has already provided many genetically modified organisms. Critics question the safety of genetically altering organisms citing unknown risks to the environment or human health.
12.1 Check Up
- Using what you know about DNA and genetics, explain how it is possible for a human gene—such as the gene for growth hormone or insulin—to be expressed in a bacterial cell from gene transfer.
- Create a concept map relating recombinant DNA to the following terms: blunt ends, ligase enzyme, plasmids, restriction enzymes, restriction fragments, restriction sites and sticky ends. Be sure to show the relationship each term has to recombinant DNA and DNA transformation experiments.
- Suppose a biotech company was producing transgenic E. coli with recombinant plasmids containing the human insulin gene. Explain the screening procedure one would use in order to determine which bacteria have taken up the vector plasmids containing the gene of interest.
- Create a list of risks and benefits of the use of transgenic technology.