6.1.3 Manipulating Genomes Flashcards
DEFINITION- Genomics
The branch of molecular biology concerned with structure, function, evolution and mapping of genomes
DEFINITION- Gene
A section of DNA that contains the complete sequence of bases (codon) to code for a protein
DEFINITION- Thermophilic
An extremophile that thrives in relatively high temperatures
DEFINITION- Thermus Aquaticus
A species of bacteria that can tolerate high temperatures
DEFINITION- TAQ Polymerase
A thermostable DNA polymerase named after the thermophile, Thermus Aquaticus
DEFINITION- Nucleotide
The monomer unit used to for nucleic acids, composed of a pentose sugar, phosphate group and nitrogenous base.
DEFINITION- H Bond
Weak bonds that can occur whenever theres a slightly negative molecule and slightly positive H
DEFINITION- DNA Primer
A short single stranded nucleic acid utilised by all living organisms in the initiation of DNA synthesis
DEFINITION- Amplification
A mechanism leading to multiple copies of a chromosomal region of a gene.
Role of PCR
DEFINITION- Restriction enzyme
Enzymes that chop strands of DNA into small pieces,
Endonucleases
DEFINITION- DNA ligase
An enzyme involved in DNA replication, that catalyses the formation of a phosphodiester bond.
DEFINITION- Plasmid
A genetic structure in a cell that can replicate independently of chromosomes
DEFINITION- Transgenic
An organism containing genetic material into which DNA from an unrelated organism has been artificially introduced.
Restriction enzyme Details
- Extracted from bacteria
- Endonucleases
- Cut DNA at specific points to create palindromic sequences
- Leave behind sticky ends
Restriction enzyme process
- Each enzyme is specific for a certain base sequence
- The active site on the enzyme has a specific shape
- The base sequence has a complimentary shape
- DNA sugar phosphate backbone is cut at the restriction site
- Hydrolysis breaks the backbone in different places
- Leaves a staggered cut (sticky end)
Polymerase Chain Reaction steps and Role
- Denaturation
- Annealing
- Extension
Amplifies DNA
Reaction mixture PCR
Extracted DNA
Nucleotides
DNA polymerase
Primers
Denaturation PCR
95degrees
H bonds break between DNA strands
Annealing PCR
55degrees
Primers bind to strands with H bonds as DNA polymerase cannot bind to single strands
Extension PCR
72degrees
DNA polymerase adds free nucleotides in 5 to 3 direction with complimentary base pairs
Differences between PCR and DNA replication
- Only short sequences can be replicated, not whole chromosomes
- A primer is required
- Needs to heat and cool to separate the DNA strands, bind on the primers and for DNA replicate
Applications of PCR
- Forensic Science
- Detecting mutations
- Monitoring spread of disease
Advantages of PCR
- Quicker
- Requires less equipment
- Less space
- Easier and less costly to run
- Safer
Advantages to cloning in living organisms (bacterial host)
- Longer sections of DNA can be cloned
- Less prone to mutation (taq polymerase can insert wrong base)
- Less expensive set up costs
- Less technically complex
Gel Electrophoresis Details
- DNA samples are treated with restriction enzymes to create DNA fragments
- Placed into wells at the negative end of the gel
- Gel immersed into buffer solution
- Electrodes are attached so a current can pass through over a set time (2hr)
- Position of fragments can be shown using dye that stains DNA
- Separates based on size
Why DNA is drawn towards the positive pole?
Gel Electrophoresis
DNA is negative so repelled away from the -ve electrode and attracted to the positive