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