Microbial Biotechnology Flashcards
When are Primary Metabolites produced?
- During log phase
When are Secondary Metabolites produced?
- During stationary phase
What is Genetic Engineering?
- Direct manipulation of an organism’s genome using biotechnology
Define Biotechnology
- The use of genetically engineered organisms to carry out defined chemical processes for industrial application
What are the Steps of Genetic Engineering?
- Isolate DNA containing target gene from human cells
- Amplify target gene with PCR
- Cut the PCR product with restriction enzymes
- Ligate the gene into a plasmid which has been cut with the same restriction enzymes
- Transform the plasmid into bacterial cells (eg. E. coli)
- Select cells that contain the plasmid with the inserted gene
- Grow cells in large batches – protein is produced
- Isolate / purify protein.
What is the Purpose of PCR?
What are the Basic Steps of PCR?
- PCR amplifies DNA
- Denaturation
- High temperature permits separation of DNA strands
- Annealing
- Cooler temperature allows primers to attach ends of target sequence
- Extension
- Temperature allowing thermostable polymerase to add nucleotides at the 3’ end of primers
What are the Advantages of using E. Coli as a Cloning Host?
- Well-developed genetics
- Many strains available
- Best-known prokaryote
What are the Disadvantages of using E. Coli as a Cloning Host?
- Potentially pathogenic
- Periplasm traps proteins
What are the Advantages of using Bacillus Subtilis as a Cloning Host?
- Easily transformed
- Nonpathogenic
- Naturally secretes proteins
- Endospore formation simplifies culture
What are the Disadvantages of using Bacillus Subtilis as a Cloning Host?
- Genetically unstable
- Genetics less developed than in E. Coli
What are the Advantages of using Saccharomyces Cerevisiae as a Cloning Host?
- Well-developed genetics
- Non pathogenic
- Can process mRNA and proteins
- Easy to grow
What are the Disadvantages of using Saccharomyces Cerevisiae as a Cloning Host?
- Plasmids unstable
- Will not replicate most prokaryotic plasmids
Describe the Production of Recombinant Insulin
- Peptide hormone
- Type 1 diabetes: insufficient insulin secretion
- need insulin injections
- Small protein consisting of 2 chains
- A & B, held together by disulphide bridges
- Two chains produced separately, then mixed
What is Synthetic Biology?
- Design and construction of new biological entities such as enzymes, genetic circuits and cells or the redesign of existing biological system
What is the Use and Principle of the Ames Test?
- Use: To determine if a particular chemical/drug is mutagenic/carcinogenic
- Principle: Chemicals that cause tumours in humans also induce mutations in bacterial cells
