Gene Tech & applications Flashcards
Describe how DNA is replicated in a cell
- DNA strands separate / hydrogen bonds broken;
- Parent strand acts as a template / copied / semi-conservative replication;
- Nucleotides line up by complementary base pairing; (Adenine & Thymine etc)
- Role of DNA polymerase: joins adjacent nucleotides on the developing strand via condensation and formation of phosphodiester bond;
- 5’ to 3’ direction
- Each new DNA molecule has 1 template and 1 new strand
- Formed by semi-conservative replication.
Why is the DNA heated to 95°C during PCR?
- Produce single stranded DNA
- Breaks WEAK hydrogen bonds between strands
Why do you add primers during PCR?
- Attaches to / complementary to start of the gene / end of fragment;
- Replication of base sequence from here;
- Prevents strands annealing
Explain why ‘base-pairs’ is a suitable unit for measuring the length of a piece of DNA.
- DNA = 2 chains / joined by linking of 2 bases / A with T and G with C/ purine pairs with pyrimidine;
- Bases are a constant distance apart / nucleotides occupy constant distance/
- each base-pair is same length / sugar-phosphate is a constant distance;
A deletion mutation occurs in gene 1.
Describe how a deletion mutation alters the structure of a gene
- removal of one or more bases/nucleotide;
- frameshift /(from point of mutation) base sequence change;
Describe a plasmid.
- circular DNA;
- separate from main bacterial DNA;
- contains only a few genes;
Suggest one reason why DNA replication stops in the polymerase chain reaction
- Limited number of primers/nucleotides; /
Primers / nucleotides ‘used up’. - DNA polymerase (eventually)denatures
Suggest why the restriction enzyme has cut the human DNA in many places but has cut the plasmid DNA only once.
- enzymes only cut DNA at specific base sequence/recognition site/specific point;
- sequence of bases/recognition site/specific point (on which enzyme acts)
- occurs once in plasmid and many times in human DNA;
- (max 1 if no reference to base sequence or recognition site)
Explain what is meant by a vector.
- Carrier;
- DNA/gene; (context of foreign DNA)
- Into cell/other organism/host;
Explain how modified plasmids are made by genetic engineering and how the use of markers enable bacteria containing these plasmids to be detected.
- isolate TARGET gene/DNA from another organism/mRNA from
- cell/organism;
- using restriction endonuclease/restriction enzyme/reverse transcriptase to
- get DNA;
- produce sticky ends;
- use DNA ligase to join TARGET gene to plasmid;
- also include marker gene;
- example of marker e.g. antibiotic resistance;
- add plasmid to bacteria to grow (colonies);
- (replica) plate onto medium where the marker gene is expressed;
- bacteria/colonies not killed have antibiotic resistance gene and (probably) the TARGET gene;
- bacteria/colonies expressing the marker gene have the TARGET gene as well;
mRNA may be described as a polymer. Explain why.
- Made up of many (similar) molecules/monomers/nucleotides/units
describe how genetic fingerprinting is carried out
- DNA extracted from a sample
- DNA cut /hydrolysed into segments using restriction endonuclease
- Must leave VNTRs /mini satellites intact
- DNA fragments separated by electrophoresis
- Detail of process e.g mixture put into wells on gel and electric current passed through
- Immerse gel in alkaline solution / two strands of DNA separated
- Radiocative marker
- Identified using x ray film
What is recombinant DNA
- cell having 2 or more sources of DNA
what is the function of reverse transcriptase
- carries out transcription in reverse, makes cDNA from mRNA templates
what is the function of restrictive endonuclease
- hydrolyse DNA at specific recognition (base) sequences
