3.8.4.1 Recombinant DNA technology Flashcards
What does recombinant DNA technology involve
Transfer of DNA fragments between organisms due to the universal genetic code and transcription translation mechanisms
What are the three methods of producing DNA fragments
Reverse transcriptase from mRNA restriction enzymes to cut DNA gene machine to synthesise
What enzyme converts mRNA into cDNA
Reverse transcriptase
What enzyme cuts DNA into fragments
Restriction endonuclease
How does a gene machine create DNA fragments
Synthesises DNA sequences based on known base sequences
What are the two methods of amplifying DNA fragments
In vitro PCR and in vivo culturing transformed host cells
What is the in vitro method for DNA amplification
Polymerase Chain Reaction PCR
What is the in vivo method for DNA amplification
Culturing transformed host cells
Why are promoter and terminator regions added to DNA fragments
Ensure correct transcription and translation in host cells
What is the role of restriction enzymes and ligases in recombinant DNA
Restriction enzymes cut DNA ligases join DNA into vectors
How are host cells transformed with recombinant DNA
By inserting vectors containing the DNA fragment
How can genetically modified GM cells be detected
Using marker genes in the recombinant DNA
Why are marker genes used
To identify cells that have successfully taken up the recombinant DNA
What are the ethical considerations of recombinant DNA technology
Balancing humanitarian benefits against environmentalist and anti-globalisation concerns
How is recombinant DNA technology related to gene therapy
It allows insertion of functional genes to treat genetic disorders
What are the ethical issues of recombinant DNA in agriculture industry and medicine
Concerns over unnatural modifications animal welfare and human safety
What are the financial issues of recombinant DNA in agriculture industry and medicine
High development costs patenting by companies creating monopolies
What are the social issues of recombinant DNA in agriculture industry and medicine
Access inequality reliance on biotech companies potential for job losses
Describe the role of Restriction endonuclease
cut plasmid/vector;
Describe the role of ligase
joins gene/DNA to plasmid/vector;
Suggest two features of the structure of different proteins that enable them
to be separated by gel electrophoresis.
- Mass/number of amino acids/polypeptides;
Accept weight for mass
Ignore density/size
Accept length of polypeptide/amino acid chain
Accept primary structure /sequence of amino acids.
Accept tertiary structure
- Charge;
- R groups (differ);
Describe and explain how the polymerase chain reaction (PCR) is used to
amplify a DNA fragment.
- (Requires DNA fragment) DNA polymerase, (DNA) nucleotides and
primers; - Heat to 95 °C to break hydrogen bonds (and separate strands);
Accept temperature in range 90 to 95 °C. - Reduce temperature so primers bind to DNA/strands;
Accept temperature in range 40 to 65 °C. - Increase temperature, DNA polymerase joins nucleotides (and repeat
method);
Accept Taq polymerase for DNA polymerase.
Accept temperature in range 70 to 75 °C.
Describe how the insulin gene is inserted into bacterial cells.
cut open vector/plasmid with a restriction enzyme
join insulin gene with vector/plasmid using (DNA) ligase
mix (recombinant) plasmids with bacteria
use ice-cold calcium chloride solution to allow bacteria to take up plasmid
OR
use electroporation to allow bacteria to take up plasmid
Describe how the researchers can obtain DNA from mRNA.
reverse transcriptase
produces complementary DNA OR cDNA from mRNA
Describe how DNA can be visualised after electrophoresis has been completed.
radioactive labels/tags
fluorescent labels/tags
UV light/radiation
visible stain
Explain how the ladder enables the sizes of the different DNA fragments to be determined.
ladder has DNA fragments of known sizes/lengths
so can compare position of fragments (to these known sizes).
Suggest how the herbicide resistance gene was used as a genetic marker.
insert the herbicide resistance gene next to the gene for the toxic protein
apply herbicide to transformed plant cells
survivors have the gene for the toxic protein
allows identification of insect-resistant soybean plants
