Appendix Flashcards
Erwin Chargaff
1949, discovered Chargaff’s Rule. Ratio of bases constant within a species. A always occurs in the same amount as T, and the same applies to G and C. From this evidence, Watson and Crick suggested that each rung of the helix is composed of a base pair.
Hershey and Chase
1952, blender experiment, discovered that DNA is the genetic material, not protein. Used a bacteriophage and labelled either protein coat with sulfure or DNA with phosphorus. Viruses and bacteria were agitated to separate them. Found that bacteria were only radioactive when DNA was labelled.
Rosalind Franklin
- Used X-ray crystallography to show that DNA is a double helix
Watson and Crick
- Stole Franklin’s picture to deduce structure of DNA
Gel electrophoresis procedure
- PCR used to amplify available amount of DNA
- Restriction enzymes used to cut DNA into fragments
- DNA is separated by placing fragments into agarose gel. DNA attracted to positive terminal. Smaller fragments move faster.
Regions used for DNA profiling
Coding regions are identical and useless. Variations occur in non-coding regions, called variable length tandem repeats.
Name of fragments in DNA profiling
Restriction Fragment Length Polymorphisms
Limitations of DNA profiling
Contamination, denatured DNA, not enough DNA
Natural examples of clones
Monozygous twins, asexual reproduction
Benefit of cloning
Can be difficult to produce genetically modified organisms, easier to make 1 then clone
2 methods of cloning
- Splitting of embryo
2. Somatic cell nuclear transfer
Explanation of embryo fragmentation
In the early stage of development, all cells making up an embryo are totipotent, and it is possible to split an 8 cell embryo into 8 cells, which would then be implanted into surrogates to become 8 clones
Somatic cell nuclear transfer
- Donor cells are taken from the organism to be cloned and cultured in a lab
- Unfertilized egg is taken from another organism
- Egg is enucleated
- Enucleated egg is fused with the donor cell
- Fused cell is cultured until an embryo is formed
- Embryo is transplanted into a surrogate
Note: not entirely identical because mDNA comes from egg
Procedure for gene transfer
- Desired gene is isolated. mRNA is first isolated, then reverse transcriptase is used to convert it to cDNA
- Bacterial plasmid is isolated
- Plasmid is prepared using restriction endonucleases
- Gene and opened plasmid are merged in a process called ligation, using DNA ligase
- Plasmid transferred back to bacterium
PCR procedure
Polymerase chain reaction.
- Denaturation. Temperature is increased to separate DNA into single strands
- Annealing. Temperature is decreased to allow primers to bind to DNA, bracketing the target sequence
- Extension. Temperature increased slightly and enzyme taq polymerase binds to DNA to add nucleotides
Process is repeated and the amount of DNA increases exponentially
Chromatography definition
Technique used to separate mixtures based on the movement of different substances on a piece of paper by capillary action
Mobile and stationary phases in paper chromatography
Stationary: paper
Mobile: solvent, usually a mix of propanone and petroleum ether
Mobile and stationary phases in thin layer chromatography
Stationary: silica gel, aluminium oxide, or cellulose
Mobile: solvent
Retention factor
Ratio of the distance moved by a pigment to the distance moved by the solvent. Constant, value for an unknown substance can be compared to known values for identification
