Module 8 - Gene probes, genetic fingerprinting and medical diagnosis Flashcards
6 marks
Describe how genetic fingerprinting is carried out.
1 DNA extracted from sample
2 DNA cut using restriction enzymes
3 must leave VNTRS / required core sequences intact
4 DNA fragments separated using electrophoresis
5 detail of process e.g. mixture put into wells on gel and electric current passed through
6 immerse gel in alkaline solution / two strands of DNA separated
7 Cover with nylon to absorb DNA
8 DNA fixed to nylon / membrane using uv light
9 radioactive marker / probe added (which is picked up by required fragments) / complementary to VNTRS
10 (areas with probe) identified using X-ray film / autoradiography
Explain genetic fingerprinting
- Collect the DNA and use PCR to amplify it.
- Restriction endonucleases are used to hydrolyse the DNA.
- Use gel electrophoresis to separate the DNA fragments.
- The DNA fragments are separated based on charge with smaller fragments of DNA moving faster and further.
- The DNA should be treated with an alkali to make it single stranded.
- The DNA probes can then be added and would hybridise with the DNA fragments.
- Wash away excess probes which did not bind.
- The probe would have fluorescence or a radioactive tracer bound to it.
- Use UV light or an audioradiography to determine if the gene/allele/VNTR is present.
6 marks
Describe how the technique of genetic fingerprinting is carried out and explain how
it can be used to identify a person, such as Saddam Hussein.
- DNA is cut
- using restriction enzyme
- electrophoresis
- separates according to length / mass / size
- DNA made single-stranded
- transfer to membrane / Southern blotting
- apply probe
- radioactive / single stranded / detected on film / fluorescent
- reference to VNTRs
- pattern unique to every individual
1 mark
What is a DNA probe?
- Short, single strand of DNA
- Bases complementary with DNA
1 mark
The DNA on the nylon membrane is treated to form single strands. Explain why.
So DNA probe binds/anneals
2 marks
The soybean cells were screened for the presence of the P34 protein. This process involved the use of gel electrophoresis to separate proteins extracted from soybean cells.
Suggest two features of the structure of different proteins that enable them to be separated by gel electrophoresis.
- Mass/number of amino acids/polypeptides
- Charge
- R groups differ
3 marks
Explain why it takes longer to obtain a genetic fingerprint if the sample is
(i) very small
(ii) contaminated
- i) PCR is needed
- ii) other DNA present
- Need to identify ‘required’ DNA from rest
2 maarks
The length of the replicated DNA fragment is different for each species.
Explain why this is important in identifying the species involved.
- Primer has different DNA sequence
- DNA specific / complementary base-pairing
2 marks
Name two techniques the scientists may have used when analysing viral DNA to determine that the viruses were closely related.
- The polymerase chain reaction
- Genetic fingerprinting
- Gel electrophoresis
- Genome sequencing
BHA
Genetic fingerprinting
13 marks
- DNA extracted from sample
- DNA amplified using PCR
- DNA cut into many different lengths using restriction enzymes
- VNTRs used as more unique than genes
- Care taken to not cut through desired VNTR section (mini satellites)
- DNA fragments placed into agar gel at negative end of circuit
- Alkaline buffer added
- Electrophoresis- DNA seperated due to charge and size
-DNA negative so moved to postive electrode
-Smaller fragments more faster and further - DNA converted to single strand and complementary DNA probe added with radioactive isotope or fluoresence- DNA hybrid formed
- Wash to remove unbound DNA probes
- Transferred to nylon sheet and fixed with UV light- southern blotting
- If radioactive isotope used on probe- transfer to X-ray film and use X-rays to identify DNA hybrids
- Must compare to known sample. If band in same horizontal position then it is the same VNTR present
2 marks
Explain how genetic fingerprinting allows scientists to identify the father of a particular panda cub
- All bands in cub which don’t come from mother
- Must be in father’s DNA fingerprint
1 mark
When pandas are bred in zoos, it is important to ensure only unrelated pandas breed. Suggest how genetic fingerprints might be used to do this.
Select pairs with dissimilar DNA fingerprints
2 marks
What is a DNA probe?
- Short single strand of DNA
- Bases complementary with DNA/allele
2 marks
DNA probes are used to detect specific base sequences of DNA. Describe how the DNA is broken down into smaller fragments.
genetic fingerprinting
- Restriction enzymes
- Cuts DNA at specific base sequence
OR
Breaks phosphodiester bonds
OR
Cuts DNA at restriction site
1 mark
The DNA on the nylon membrane is treated to form single strands. Explain why
So DNA probe binds/anneals
2 marks
Short tandem repeats (STRs) are short sequences of DNA, usually 2 to 7 base pairs. STRs are repeated a number of times, one after another. For example, the STR D5S818 is made up of AGAT repeated 7 to 16 times.
STRs are found throughout the whole genome.
The repeated sequences in STRs are common to all humans. Due to variation in the number of repeats, STRs can be used in genetic fingerprinting. Describe how STRs could be removed from a sample of DNA.
- Restriction enzymes
- Cut DNA at specific base sequences
2 marks
Genetic fingerprinting using STRs requires amplification of the STRs using the polymerase chain reaction (PCR). The short base sequences either side of a specific STR are known.
Explain the importance of knowing these base sequences in PCR.
- For primers
- To produce a complementary base sequence
- Primers mark beginings and ends of parts of DNA for attachement of enzymes (DNA taq polymerase) and DNA nucleotides
2 marks
During genetic fingerprinting, the different STRs are separated by gel electrophoresis.
Give two features of STRs which enable them to be separated by gel electrophoresis.
- Number of nucleotides
OR
Length/mass - Negative charge
