genome projects and gene technology 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.
Describe and explain how the polymerase chain reaction (PCR) is used to amplify a DNA fragment.
- (Requires DNA fragment) (Taq) DNA polymerase, (DNA) nucleotides and primers;
- Heat to 95°C to break hydrogen bonds (and separate stands);
- Reduce temperature (40-65°C) so primers bind to DNA/strands;
- Increase temperature (70-75°C), DNA polymerase joins nucleotides (and repeat method);
Why is the DNA heat 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;
Name one mutagenic agent.
- high energy radiation /ionising particles e.g. named particles/α, β, γ & X-rays;
- benzene;
- x rays/cosmic rays;
- uv (light);
- carcinogen / named carcinogen;
- mustard gas / phenols / tar (qualified);
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 the main stages in the copying, cutting and separation of the DNA.
- heat DNA to 95°C / 90°C;
- strands separate;
- cool so that primers bind to DNA;
- add DNA polymerase/nucleotides;
- use of restriction enzymes to cut DNA at specific base sequence/ breaks phosphodiester bonds
- use of electric current and agar/gel;
- shorter fragments move further;
Describe the polymerase chain reaction.
- Heat DNA;
- Breaks hydrogen bonds/separates strands;
- Add primers;
- Add nucleotides;
- Cool;
- (to allow) binding of nucleotides/primers;
- DNA polymerase;
- Role of (DNA) polymerase;
- Repeat cycle many times;
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)
Describe how the bacteria containing the insulin gene are used to obtain sufficient insulin for commercial use.
- use of fermenters;
- provides nutrients plus suitable conditions for optimum growth/named
- environmental factor;
- reproduction of bacteria;
- insulin accumulates and is extracted;
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;
