Biotechnology and Genetic Engineering - Genetic engineering (Insulin production)
Review the key concepts, formulae, and examples before starting your quiz.
🔑Concepts
Genetic engineering is the process of altering the genome of an organism by introducing a gene from another organism to achieve a desired phenotype, such as the production of human insulin by .
Restriction Endonucleases: These enzymes act as 'molecular scissors' that cut DNA at specific palindromic sequences. They often create 'sticky ends'—short, single-stranded overhangs that allow for complementary base pairing with a foreign gene.
DNA Ligase: This enzyme acts as 'molecular glue,' forming phosphodiester bonds to join the sugar-phosphate backbones of the insulin gene and the plasmid, resulting in recombinant DNA.
Plasmids as Vectors: Small, circular loops of DNA found in bacteria that are used to transport the human insulin gene into the host bacterial cell.
Reverse Transcriptase: Used to synthesize complementary DNA () from insulin extracted from pancreatic -cells. This ensures the gene does not contain introns, which bacteria cannot process.
Selection and Fermentation: Transformed bacteria are identified using marker genes (like antibiotic resistance) and then grown in large-scale fermenters under controlled conditions (, , and temperature at ) to mass-produce the protein.
The produced insulin is chemically identical to human insulin, reducing the risk of allergic reactions compared to bovine or porcine insulin.
📐Formulae
💡Examples
Problem 1:
A scientist extracts for insulin from a human cell. Why is it necessary to use reverse transcriptase to produce before inserting it into a bacterial plasmid?
Solution:
Bacteria are prokaryotes and lack the necessary machinery (spliceosomes) to remove introns from eukaryotic pre-. By using reverse transcriptase to create , the scientist ensures the gene contains only exons (coding sequences), allowing the bacteria to translate the sequence correctly into the insulin protein.
Explanation:
Eukaryotic genes contain non-coding regions called introns. Prokaryotic organisms like cannot excise these; therefore, the 'finished' (with introns already removed) must be converted back to DNA () to be functional in a bacterial host.
Problem 2:
Explain the importance of using the same restriction endonuclease on both the human DNA fragment and the bacterial plasmid.
Solution:
Using the same restriction enzyme ensures that both the gene and the plasmid have complementary 'sticky ends'. This allows -bonds to form between matching base pairs ( with , with ), facilitating the action of DNA ligase.
Explanation:
If different enzymes were used, the overhanging sequences would not match, and the human gene would not integrate into the circular plasmid to form recombinant DNA.