21.1 DNA Profiling Flashcards
Chapter 21 - Manipulating Genomes
What is a genome?
All the genetic material of an organism.
What is an exon?
Regions of DNA that code for proteins.
What is an Intron?
Regions of non-coding DNA or RNA.
What is a telomere?
Structure at the end of a chromosome.
What is a centromere?
Region at which two chromatids are held together.
What is satellite DNA?
Short sequences of DNA that are repeated many times within the introns, telomeres, and centromeres.
What is a mini-satellite?
A sequence of 20-50 base pairs repeated 50+ times (aka variable number tandem repeats - VNTRs)
What is a micro-satellite?
A small region of 2-4 bases repeated 5-15 times (aka short tandem repeats -STRs)
What is DNA profiling?
Producing an image of the patterns in the non-coding DNA of an individual.
What is the first stage in the process of producing a DNA profile?
Extracting the DNA.
Where must the DNA be extracted from?
A tissue sample.
What technique or process is used when extracting DNA?
Polymerase Chain Reaction (PCR).
What is the polymerase chain reaction (PCR)?
A process by which a small sample of DNA can be amplified using specific enzymes and temperature changes.
Summarise in simple words, step 1 of producing a DNA profile.
DNA is extracted from the sample.
What is the second stage in the process of producing a DNA profile?
Digesting the sample.
What enzyme is used when digesting the sample?
Restriction endonucleases.
What is restriction endonucleases?
Enzymes that chop a DNA strand into lots of small pieces.
What is the restriction site or recognition site?
A specific nucleotide sequence the different restriction endonucleases cut DNA at.
How many cuts do all restriction endonucleases make?
Two, once through each strand of the DNA double helix.
What do restriction endonucleases give scientists the ability to do?
Cut DNA strands at defined points in the introns, leaving repeat units or satellites intact and a mixture of mini- and microsatellite regions intact at the end.
Summarise in simple words, step 2 of producing a DNA profile.
Restriction endonucleases cut the DNA into fragments.
What is the third stage in the process of producing a DNA profile?
Separating the DNA fragments.
What technique or process is used to separate the DNA fragments?
Electrophoresis.
What is electrophoresis?
A type of chromatography that relies on the way charged particles move through a gel under the influence of an electrical current. Used to separate nucleic acid fragments.
Why is the gel, in step 3 of producing a DNA profile, immersed in an alkaline buffer after running in electrophoresis?
Denature DNA fragments causing strands to separate into single strands and expose the bases.
In electrophoresis, why is the agarose gel placed into a buffering solution?
To maintain and prevent a decrease in pH as pH can denature enzymes.
What is the purpose of a DNA ladder?
Compare fragments of DNA to known values.
What is a DNA ladder?
A set of standards that are used to identify the approximate sizes of a molecule run on gel in electrophoresis.
What does the rate of movement depend on within an agarose gel?
The length of DNA fragments.
Small fragments with less base pairs move faster.
What is southern blotting?
Transferring separated strands to nitrocellulose paper or nylon membrane placed over the gel. The membrane or paper is covered with several sheets of dry absorbent paper to draw the alkaline solution containing DNA up by capillary action. Single-stranded DNA is transferred to the paper or membrane as they are unable to pass through and is then fixed with UV light or heating to 80°C.
Summarise in simple words, step 3 of producing a DNA profile.
Fragments are separated using gel electrophoresis.
DNA fragments are transferred from the gel to a nylon membrane by the process of southern blotting.
What is the fourth stage in the process of producing a DNA profile?
Hybridisation.
What is hybridisation?
The addition of fluorescent or radioactive probes to DNA fragments via complementary base pairing.
What do DNA probes identify?
The microsatellite regions that are more varied than the larger minisatellite regions.
Summarise in simple words, step 4 of producing a DNA profile.
DNA probes are added to label the fragments. These radioactive probes attach to specific fragments.
What is the final stage in the process of producing a DNA profile?
Seeing the evidence.
How would you see the evidence if radioactive labels were added to DNA probes?
X-ray images are taken of the paper/membrane.
How would you see the evidence if fluorescent labels were added to DNA probes?
Paper/membrane is placed under UV light so the fluorescent tags glow.
What are the ingredients needed to carry out the Polymerase Chain Reaction (PCR)?
- A sample of DNA.
- Free nucleotides (A,T,C,G).
- DNA polymerase (Enzyme).
- Primers (Complimentary to the beginning of the target sequence, like a starting point).
What is a Polymerase Chain Reaction (PCR) machine also called?
Thermal Cycler.
What is the first stage when carrying out a Polymerase Chain Reaction (PCR)?
Denaturation.
What happens in the first stage of a Polymerase Chain Reaction (PCR)?
Temperature set to 95°C causing DNA stands to separate by the breaking of the hydrogen bonds.
What is the second stage when carrying out a Polymerase Chain Reaction (PCR)?
Annealing.
What happens in the second stage of a Polymerase Chain Reaction (PCR)?
Temperature decreased to 55°C to allow primers to attach (anneal) to complimentary ends of the DNA template strand.
What is the third stage when carrying out a Polymerase Chain Reaction (PCR)?
Extension.
What happens in the third stage of a Polymerase Chain Reaction (PCR)?
Temperature increased to 72°C to allow Taq polymerase to add complimentary bases to the template strand. (like its extending from the primers by adding bases).
What are some uses of DNA profiling?
- Forensic science (Crime scenes)
- Proving paternity of a child (prove or disprove family relationships)
- Identifying the species to which an organism belongs to.
- Demonstrate evolutionary relationships between different species.
- Identifying patients who are at risk of developing particular diseases (eg. cancers and heart diseases)
