CS262 Discussion Section 2. Today’s agenda. Overview of some more biology underlying DNA sequences. Problem Set difficulties. Molecular scissors. Enzymes, naturally occurring in bacteria, that cut DNA at very specific places. e.g, BamHI cuts recognition sequence GGATCC between GG.
Today’s agenda • Overview of some more biology underlying DNA sequences. • Problem Set difficulties
Molecular scissors • Enzymes, naturally occurring in bacteria, that cut DNA at very specific places. • e.g, BamHI cuts recognition sequence GGATCC between GG. • Another restriction enzyme will have a different recognition sequence and will make its cuts between different pairs of bases.
Viruses: Making a living by hijacking cells • A virus is like a thief who arrives at a factory he intends to rob possessing only two things – the tools to get inside, and some software that will make the factory turn out items he can use. T4 bacteriophage is a virus that looks like an alien landing pod. With its six legs, the bacteriophage attaches to the surface of the much larger bacteria Escherichia coli (E. coli).
Once attached, the bacteriophage injects DNA into the bacterium. The DNA instructs the bacterium to produce masses of new viruses. • So many are produced, that the E. coli bursts.
Restriction-modification systems • A modification methyl-transferase that recognizes a specific DNA sequence, and methylates particular bases in that sequence. • A restriction endonuclease that recognizes the same sequence, and if the site is not methylated, cleaves the DNA.
Biotechnology • The use of living organisms to create products. • Human Growth Hormone (HGH) • Secreted by pituitary gland • A faulty pituitary gland can leave people abnormally short. • Previously, HGH was laboriously extracted from the pituitaries of a dead human. • Unsafe and produces too little HGH
Biotechnology: Produce synthetic HGH • Isolate gene for HGH from human cells and snip it out. • Insert into E.coli. • E.coli will start transcribing and translating the gene. Grow the bacteria in millions. • Result: Biotech firms manufacture HGH and ship to pharmacies worldwide.
How do you cut DNA to get a human gene out of the human genome? • Use restriction enzymes. • How do you get this gene coding for protein inside a group of bacterial mini-factories?
Plasmids • Plasmids are extrachromosomal rings of bacterial DNA that can be as little as 1000 bp in length. • They can replicate independently of the bacterial chromosome, and they can move into bacterial cells. • Bacteria are capable of taking up DNA from their surroundings, after which this DNA will code for proteins inside the bacterial cell. (Transformation)
Definitions • Recombinant DNA: Two or more segments of DNA that have been combined by humans into a sequence that does not exist in nature. • Cloning: Making an exact genetic copy. A clone is one of the exact genetic copies. • Cloning vector: Self-replicating agents that serve as vehicles to transfer and replicate genetic material.
The wider world of biotechnology • Cloning can involve not just genes but whole organisms. • Humans have actually been making clones for centuries, eg, ‘cuttings’ taken from plants. • Biotechnology has expanded the range of what can be cloned.
Scotland, 1997 • Dolly, the sheep, cloned by Ian Wilmut and colleagues. • Reproductive cloning: Cloning intended to produce genetically identical animals.
How was Dolly cloned? • Dolly is an exact genetic replica of another sheep.