8.4 Gene Technologies Flashcards
Why does recombinant DNA technology work?
Genetic code is universal
Transcription/translation occur by the same mechanism and result in the same amino acid sequence across organisms
What is meant by recombinant DNA technology?
Transfer of DNA fragments from one organism to the other
Summarise the process of using reverse transcriptase to produce DNA fragments.
mRNA complementary to the target gene is used as a template
Mixed with free nucleotides which match up to their base pairs
Reverse transcriptase forms sugar-phosphate backbone to create cDNA
What is cDNA?
Complementary DNA
Summarise the process of using enzymes to produce DNA fragments.
Restriction endonucleases cut DNA at specific sequences
Different REs cut at different points but one RE will always cut at the same sequence
Using particular REs allows you to cut out a certain gene of interest
What two ways can DNA fragments be amplified?
In vitro
In vivo
What is in vitro?
Polymerase Chain Reaction (PCR)
What is in vivo?
Using host cells
Describe the reaction mixture in the first stage of PCR.
Contains the DNA fragment to be amplified
Primers that are complementary to the start of the fragment
Free nucleotides to match up to exposed bases
DNA polymerase to create the new DNA
Summarise the process of amplifying DNA fragments using PCR.
Heated to break apart the DNA strands
Cooled to allow primers to bind
Heated again to activate DNA polymerase and allow free nucleotides to join
New DNA acts as template for next cycle
Summarise the process of inserting a DNA fragment into a vector.
A plasmid is used as the vector
Its cut using the same restriction enzymes as the DNA so that the ends are complementary
DNA ligase joins the fragment and plasmid together
What is a plasmid?
Circular DNA from bacteria
Summarise the process of inserting a vector into a host cell.
The host cells are mixed with vectors in an ice-cold solution
Heat shocked to encourage cells to take up vectors
The cells can be grown and the DNA fragment will be cloned
What is the host cell?
Bacteria
What does in vivo DNA amplification do?
Transform cells
Summarise the process of identifying transformed cells.
Marker genes
UV light can be used to identify which cells have taken up vector and which haven’t
How can DNA probes be used to locate specific alleles?
Designed so that its sequence is complementary to the allele to find
Probes are labelled, amplified using PCR and added to sample of single stranded DNA
Probe will bind if allele is present
Give some applications of DNA probes.
Screen someone’s DNA for a heritable health condition
Identify a gene for use in genetic engineering
Predict a patient’s response to a drug
What is the purpose of DNA hybridisation?
To measure the degree of difference between two strands of DNA
Can be used to compare DNA to see if a certain gene is present
Summarise the process of DNA hybridisation.
DNA strand is labelled and mixed with an unlabelled comparison strand
The more similar the strands, the more strongly they bind and more energy required to break strands apart
What are the benefits of genetic profiling?
Can identify heritable diseases early
Treatment can be personalised
What is genetic fingerprinting?
A technique used to compare two DNA samples and determine whether they came from the same individual
What is meant by VNTRs?
Variable Number Tandem Repeats
The non-coding regions contained in every organism’s genome
How does genetic fingerprinting work?
The probability of two individuals having the same VNTRs is low so these areas are compared to see if two DNA samples came from the same organism
