genetic engineering what is genetic engineering? genetic engineering = inserting a foreign gene of...
TRANSCRIPT
Genetic Genetic EngineeringEngineering
What is Genetic Engineering?
• Genetic Engineering = inserting a foreign gene of interest into a host to transcribe and translate a particular protein.
• Ex. Inserting the human insulin gene into bacteria to mass produce it.
Image taken without permission from http://www.medicalprogress.org/uploads/images/insulin%20inject%20WSU%20210.jpg
General Steps
• Obtain the gene of interest (ex. insulin gene)
• Insert the gene into the host (ex. bacteria)
• Allow the host to multiply and express the foreign gene– get your desired protein!– Get lots of cells that can make the protein = clones
The Big Picture
• The inserted gene is transcribed and translated using the RNA Polymerase, ribosomes and other resources in the cell
Plasmids
• Circular DNA
• Extrachromosomal – NOT part of the E. coli genome– “extra” DNA
• Contain a few non-essential genes
• Can give the bacteria additional “traits”– Depends on the genes on the plasmid
• Can be exchanged between bacteria
chromosomal DNA
plasmids
Recombinant plasmids
• Plasmids can be modified in biological labs
• Modified plasmid = Recombinant plasmid
• Plasmids can be used as cloning vectors to get the recombinant plasmid into E. coli– Cloning vectors = way to get the gene of
interest into the host
Transformation
• Process in which foreign DNA is physically inserted into host E. coli cells.
• E. coli that contains recombinant plasmid = Transformed cell Image taken with out permission from
http://summerschool.at/static/irismaria.schoenbrunner/images/transformation.png
Transformation Steps
• Recombinant plasmids and host E. coli are mixed together
• CaCl2 is added
– The Ca2+ ions neutralize the negative charges on plasmid DNA
– Help plasmid enter the membrane
Image taken without permission fromTransformation Animation. Available at http://www.dnai.org/b/index.html
Transformation Steps
• Heat Shock– By rapidly changing the temperature of the
solution, temporary pores are opened in the membrane
– Creates an opening for the plasmids to enter the E. coli
Transformation
• Transformation is not 100% successful
• After transformation– Some cells will contain plasmid = transformed– Some cells won’t contain plasmid = untransformed
• In a later step, you will determine which cells were transformed
E. Coli as a host
• E. coli is a good host because:– Reproduce quickly (once every 20 minutes)– Nonpathogenic (the strain we use is not
harmful)– Genome fully characterized (all genes have
been sequenced)