Biotechnology Flashcards
Describe how genetic fingerprinting may be carried out on a a sample of DNA
DNA is cut; using restriction enzyme; Use electrophoresis; Separates according to length/mass; Southern blotting/transfer to (nylon) membrane; Make single-stranded; Apply probe; Radioactive or fluorescent; Reference to tandem repeats/VNTRs/minisatellites;Autoradiography/eq;
List Biotechnology Techniques related to manipulating DNA
- Restriction Enzymes
- DNA Ligase
- Reverse Transcriptase
List biotechnology techniques related to analysing DNA
- PCR
- DNA Sequencing
- Restriction Mapping
- Electrophoresis
- Southern Blot
- Genetic Fingerprinting
List biotechnology techniques related to cloning DNA
- Vectors
- Transformation
- Marker Genes
What do restriction enzymes do?
Cut DNA at specific sites.
What are sticky ends, as created by restriction enzymes?
Sticky ends are ends with exposed bases that can anneal onto other sticky ends that have been cut with the same enzyme)
What are blunt ends, as created by restriction enzymes?
Ends with no exposed bases
What is the place where restriction enzymes are cut called?
Recognition sites
What is DNA Ligase?
An enzyme that joins two nucleotides together. Often used to join together complementary restriction fragments
What is Reverse Transcriptase?
an enzyme which synthesised DNA from an RNA template
Where does Reverse Transcriptase come from?
Retroviruses
What is the kind of DNA created by Reverse Transcriptase from mature mRNA called?
cDNA (Complementary DNA)
How can Reverse Transcriptase be used in biotechnology?
- Make genes without introns to splice into plasmids
- Makes a stable copy of a gene (less readily broken down than RNA)
- Makes genes easier to find
Explain how the strands of DNA are separated during the PCR
by heating to break the H-bonds (between complementary bases)
in PCR why are primer required
to allow the DNA polymerase to attach
start addition of nucleotides
mark start and end of sequence to be copied
prevents strands re-joining
In PCR two different primers are required, why?
because the sequences at the ends of the target sequence are different or
one is at the beginning and one at the end;
Describe how a gene can be isolated from human DNA.
Using restriction endonuclease/enzymes to cut DNA at a specific place
Why is modified deoxy nucleotide added to test tubes during Sanger Sequencing?
To stop further synthesis of DNA, as it cannot form phosphodiester bonds
How is DNA sequencing read on the electrophoresis gel?
From bottom to top
What is a restriction map?
A diagram of a piece of DNA marked with the location of sites where it is cut by restriction enzymes
Explain Southern Blotting
- Hydrogen bonds broken between DNA bases
- Nylon and stack of paper towels weighted down on top of gel. Negative DNA drawn to positive nylon
- Nylon removed and DNA fixed by UV, then mixed with probes
- Probes anneal by complementary base pairing
- Visualised by fluorescent light or autoradiography
What is a vector?
A length of DNA that carrier the desired gene into a host cell
What are marker genes?
Genes used to find which cells have taken up the hybrid vector
What are the two types of marker genes in a Vector?
- Antibiotic Resistance Gene
- GFP
Why do we need an antibiotic resistance marker gene on top of the GFP gene?
So after the gene transformation, the cells can be cultured in that antibiotic to determine which cells have taken up the plasmid
Give two characteristic features of stem cells.
Will replace themselves/keep dividing/replicate
Undifferentiated/can differentiate/develop into other
cells/totipotent/multipotent/pluripotent
