DNA Flashcards
What is DNA?
Deoxyribonucleic Acid
- a long linear ‘polymer’ composed of small ‘monomers’ known as nucleotides.
What is a nucleotide composed of?
- A sugar (ribose)
- A Phosphate group (i.e. PO4)
- One of four bases
What are the four bases that could make a nucleotide?
- Adenine (A)
- Thymine (T)
- Guanine (G)
- Cytosine (C)
What are the specific base pairs that are made?
hint only certain ones pair with each other
- Adenine & Thymine (A-T)
- Guanine & Cytosine (G-C)
What is viral DNA?
Inserted by a virus over time but no longer has a purpose or function - however it is still replicated with cell division & passed on to offspring.
May be very unique to individuals because their ancestors will never have been exposed to the exact same virus.
What is VNTR?
Variable Number Tandem Repeats - refers to sequences up to several hundred base pairs that repeat many times.
What is STR?
Short Tandem Repeats refers to the regions in VNTR where repeating sequences is 3-7 base pairs in length & may repeat less than 30-40 times making them better suited for PCR amplification.
What is RFLP?
Restriction Fragment Length Polymorphisms
What are Restriction Sequences?
Restriction enzymes from bacteria used to ‘cut’ DNA at specific base pair sequences known as ‘Restriction Sequences’
What does Polymorphism mean in RFLP?
Single long DNA strands may be ‘cut’ into several restriction fragments with different lengths and composition.
Describe the process of RFLP
1) DNA is ‘cut’ at specific base pairs (restriction sequences)
2) DNA cut into several ‘restriction fragments’ of different lengths & compositions (polymorphism)
3) Agarose Gel, samples inserted into wells within the gel, buffer at top & bottom
4) Current turned on, fragments separate by size throughout gel
5) gel removed & stained; smaller fragments @ bottom
6) DNA extracted to nitrocellulose sheet, labelled with p32, exposed against xray to produce autoradiogram
* * produces band sequences**
What does RFLP analysis require?
- Large amounts of high quality DNA
- Sophisticated equipment
- Highly trained technicians
- Radioactive isotopes (creates waste)
- Fairly long time (several days)
** best evidence for differentiating individuals **
What is PCR?
Polymerase Chain Reaction
Describe Polymerase Chain Reaction
- Cellular DNA replication
1) unwind double helix to form two strands
2) Attach ‘RNA primers’
3) DNA polymerase attaches primer, begins to add nucleotides to form new strand of DNA
4) Repeated until entire chromosome is replicated
What steps are involved in Polymerase Chain Reaction?
1) DNA is isolated from cell (only 1 copy needed)
2) DNA is added to mixture of
* Nucleotides, DNA primers, Polymerase enzyme, ATP & other ‘stuff’
3) Mixture is placed into Thermocycler (heats & cools in repeating sequence
4) heated to separate 2 strands of DNA
5) cooled to allow primers & polymerase to DNA strands
6) heated so polymerase replicates DNA strand
7) cooled to allow new DNA to stabilize
