Immunity, Infection & Forensics Flashcards
When DNA is read what does it code for?
For a specific proteins
In a eukaryotic cell when can gene expression be regulated?
After transcription and before translation.
What does post transcriptional modification involve?
Involves modifying mRNA into mature mRNA
In eukaryotes what does the process of transcription result in?
The synthesis of pre-mRNA.
Why is Prost-transcriptional regulation required ?
Needed because eukaryotic genes contains introns and extrons
What are Exons?
Sections of DNA that code for proteins
What are Introns?
Sections of DNA that do not code for protein
What must be removed before the pre-mRNA can be synthesised into polypeptides?
Introns must be removed
Failure to remove introns would result in?
An incorrect sequence and as a direct consequence to the wrong person would get made during translation
What is splicing ?
The process in which introns are removed from pre-mRNA and the exons are joined together to form mature mRNA
What is the process of splicing catalysed by?
Enzyme - RNA complex - spliceosome
Explain why it is difficult to identify the true number of genes that are in the human genome
Primary mRNA cans be spliced and combined in many different ways.
This allows for more protein to be produced than the number of genes in human genome.
Explain the advantage of being able to modify the mRNA to produce different proteins.
Being able to modify the mRNA allows an organism to produce alternative proteins without needing an entirely new gene to code for it.
This ensures that the organism’s genome is not too large.
Describe some of the modifications that a polypeptide chain undergoes before becoming a functional protein.
modifications include:
- cleaving the chain to form the functional protein
- adding carbohydrate or phosphate group to alter the structure or destination of the protein
- attaching lipids t anchor the proteins in a cell membrane
- degrading the protein chain once it has served its purpose
Explain why changes to the polypeptide are necessary?
changes to the polypeptide determine the poteins structure and function, or determnine where the protein is transported to.
without these modifications, the proetin may not function properly.
Does the process of splicing only involve the removal of introns?
No it can also consist of removal of some exons
No it can also consist of removal of some exons
Different combinations of mature mRNA will be formed
What can one gene produce ?
Many different mRNAs hence different proteins
How are humans able to produce more over 100,000 proteins from only 20,000 genes?
Due to alternative splicing
Explain the purpose of of PCR
To produce large quantities of ‘cloned’ DNA from very small samples.
Large quantities are needed for effective analysis .
small quantities are often unusable.
Describe how the polymerase chain reactions work
- DNA is heated to 96 to denature the strands and provide single straded templates for replication
- The reaction is cooled to 60 to allow primers (short lengths of DNA to which free nucleotides attach to) to aneal to the single strand of DNA
- The reations is heated to 72 so that the Taq Polymerase can work at an optimum condition and rapidly extend the nucleotide chain from the primer
- This process is repeated 25-35 times and the amount of DNA icreases exponentially with each repeat.
Describe 3 situations where only very small DNA samples may be available for sampling and PCR could be used
forensic sample taken at the scene ofthe crime - hair, blood, semen
Archaeological samples from human remains
samples taken from the remains of prehistoric organisms preserved in ice, mummified, preserved in amber, tar pits
After two cycles of replication, 4 copies of the double-stranded DNA exists. Calculte how much a DNA sample will have inreased after a. 10 cycles b. 25 cycles
a. 10- cycles = 1024
b. 25 cycles = 33 554 432
The risk of contamination in the preparation of PCR is considerable. Explain what the effect would be of having a single molecule of unwanted DNA in the sample prior to PCR
it would be amplified along with the intended DNA sample, thereby contaminating the sample and rendering it unusable
Describe the 2 possible sources of DNA contamination in preparing a PCR sample
dirty equipments -from previous treatments → dna molecules left
DNA from the technician - dandruff
spores, viruses + bacteria in the air
Describe 2 other genetic engineering/ genetic manipulation procedures that require PCR amplification of DNA
DNA sequencing , gentic cloning
Describe the properties of the short tandem repeats that are important to the application of DNA profiling technology
STRs are non-coding nucleotide sequence (2=6 base pairs) that repeat themselves many times over.
THe human genome has numerous different STRs
This property can be used to identify the natural variation found in evey persons DNA since every person will have a different combinations of STRs of different length
Explain the role of the PCR in the process of DNA profiling
used to make copies of the STRs. Only the STR sites are amplified by the PCR becausse the primers are used to initiate the PCR are very specific
Describe the 3 main steps in DNA profiling using PCR
- Extract the DNA sample. treat the tissue with chemical and enzymes to extract DNA which is then seperated and purified
- amplify the sTR sites using PCR. Priers are used to make a large quantities of STR
- Run the fragments through a gel to separate them. The resulting pattern represents the STR size for that individual
Explain why as many as 10 STR sites are used to gain a dNA profile for forensic evidence
To ensure thata the number of STR sites, when compared, will produce a profile that is effectively unique
it provides a high degree of statistical confidence when a match occur
Explain the pupose of gel electrophoresis
Gel electrophoresis separates mixtures of molecules on the basis of size ad other physial properties
Describe the process of gel electrophoresis
- The DNA is loaded into wells at one end of a slab of agrose gel and an electric current is passed through
- The negatively charged (due to the phosphate group) DNA moves through the gel towards the positive electrode, with the smaller fragments moving faster and further
- The DNA ends up aranged in bands, with similar length strands rouping together.
Describe the two forces that control the speed at which framents pass through the gel
the frictional forces of each fragment’s size (larger fragments travel more slowly than smaller ones)
the strength of the electric field (movement is more rapid in stroner fields)
Explain why the small fragments travel through the gel the fastest
The gel is full of pores through which the fragments must pass.
Smaller fragments pass through these pores more easily with less resistance and
therefore faster than larger ones
what are the controlled variables in this practical?
Voltage applied
Thickness of agarose gel
Temperature
Time allowed for DNA to run
Restriction enzymes used
what are restriction endonucleases?
type of enzymes deried from bateria that uts into DNA fragments at speific restriction sites
What is the Southern blotting?
alkaline buffer solution is poured over salb after gel electrophoresis
use a dry nylon filter to absorb fragments of stained DNA & proue visible blobs
Summarise the process of DNA profiling
- uses restiction endonucleases to produce DNA fragments
- use gel electrophoresis to separate fragments
- label fragments wit fluorescent or radioactive gene probes
- perform Southern blotting to compare pattern of bandsr
How can one gene result in the production of several different proteins?
Post transcriptional modification of pre mRNA splices out all introns and some exons
splicesomes rejoins the fragments. Exons can be joined in vaious sequence producing different versions of functional mRNA
What are the 3 stages of PCR?
DENATURATION
ANNEALING
ELONGATION
State the difference between alternative splicing and Splicing
Introns removed in splicing
Alternative splicing some exons spliced
