D1.1 DNA replication Flashcards
(27 cards)
Who discovered the DNA structure?
Watson & Crick
What did Watson & Crick’s discovery provide evidence for?
Complementary base pairing was the key to DNA’s ability to replicate
What is DNA replication?
The process by which exact copies of DNA are created for use in reproduction, growth and tissue replacement in multicellular organisms
Why does DNA need to replicate?
- Cell division - new cells need new DNA for growth & tissue repair
- Reproduction - gametes requires DNA to pass on genetic info
What is the correct theory for DNA replication?
Semi conservative replication
How does semi conservative replication work?
- Hydrogen bonds between the bases are broken
- Free nucleotides are present in the nucleus
- Free nucleotides pair up with complementary exposed bases
- New strand linked together
- There are now 2 DNA molecules (Each contains one old and one new)
What is the function of DNA helicase?
Breaks hydrogen bonds & unwinds molecule (Unzip)
What is the function of DNA polymerase?
Form the phosphodiester bonds between nucleotides
What are the 3 different theories for DNA replication?
Dispersive
Conservative
Semi conservative
What is the evidence for semi conservative replication?
Cultured e coil bacteria in presence of a heavy nitrogen isotope
1. DNA contains nitrogens in its nitrogenous base = radioactive nitrogen will end up in DNA of the bacteria
2. Result: all bacterial DNA had nitrogen in its bases
3. Bacteria all transferred into a fresh medium (nitrogen replaced by a lighter isotope, bacteria was allowed to grow for several generations
What is gel electrophoresis used for?
Identify alleles at a few (or few dozen) loci
How does gel electrophoresis work?
Uses an electrical current to move molecules through a semi-solid medium
(molecules are usually DNA, RNA or protein - separated by size & amount of charge)
What are the key principles of gel electrophoresis?
- DNA & RNA molecules have negative electrical charge = will move towards positive electrode in an electric field
- to get fragments of appropriate size, DNA is digested with restriction endonucleases
- Samples of fragements of DNA -> loaded into wells on one end of the gel
- gel is submerged in buffer solution & electric current is run through the gel
- MUST begin at the negative pole
Smaller pieces of fragments in gel electrophoresis travel….
Further along the gel (in a given amount of time), quicker
What is used to make DNA have colour? and why?
Ethidium bromide
Binds to DNA, fluoresces in UV light
What is a limitation of using ethidium bromide to make DNA have colour?
Mutagenic - safer alternatives used
What is the ‘DNA ladder’ for?
It is a DNA size marker, by comparing the position the approximate size can be determined
What is PCR and what does it do?
Polymerase Chain Reaction
- a form of artificial DNA replication
- amplifies DNA fragments for further study or processing
What does PCR require?
- small DNA sample
- DNA polymerase
- Primers
- Nucleotides
- Thermocycler
What is a thermocycler?
Controls and changes its temperature at programmed timings to trigger the different steps in PCR
What are the stages in PCR?
Stage 1: Denature to separate DNA strands (95 degrees)
Stage 2: Annealing (joining) of primers to their complementary bases - provides a starting point for DNA polymerase
Stage 3: Synthesis of DNA (temp 72 degrees- optimum temp for DNA polymerase)
PCR is a …………….. reaction
Cyclic
What are 2 advantages of PCR?
- Fast
- Does not require living cells
What are some applications of PCR?
- detecting of oncogenes
- detecting mutations
- identify viral infections
- monitoring the effect of a viral disease
- tissue typing
- research
- forensic science
