Manipulating Genomes Flashcards
Summary
Define primer
Small sections of DNA complementary to the fragments desired
How does PCR work?
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Heat to 95 degrees - breaks hydrogen bonds between bases
Cool mixture to 50 degrees - primers bind to strand
Heat to 72 degrees - thermostable DNA polymerase lines up free nucleotides along template strand via complementary base pairing
Cycle then repeats
Action of restriction enzymes
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Restriction enzymes cut DNA at specific recognition sequences
via hydrolysis reaction
It leaves a sticky end to bind to DNA fragment
How to do electrophoresis ▪what gets poured? ▪ what's added? ▪what is mixed? ▪what's transferred? ▪whats connected? ▪how to identify?
Pour agarose gel into tray and leave to set
Add buffer solution completely submerging gel
Mix DNA fragment with loading dye
Micropipette into wells
Connect terminals of power supply and leave to run
Stain gel
What is a DNA profile?
What are 2 uses?
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Number of times a base sequence is repeated in a genome (this is used to compare DNA)
Forensics
Medical diagnosis
Define recombinant DNA
DNA formed by joining DNA from different sources
How is recombinant DNA formed? ▪cut ▪ cut ▪ pairing ▪ joining
DNA containing desired gene isolated using restriction enzymes
Same enzyme used to cut open vector DNA
Vector DNA and DNA fragment join by complementary base pairing
DNA ligase joins sugar-phosphate backbone
How to create insect repellent plants?
(simple version)▪▪▪
Plasmid used from Bt bacterium
Modified
Made to infect the plant to insert DNA into plant
Difference between somatic and germ line gene therapy
Somatic cell gene therapy alters body cells and offspring have a chance of getting disorder
Germ line cell gene therapy alters sex cells and offspring have no chance to get disorder
Disadvantages of gene therapy
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Body can identify vector as pathogen, stimulate immune response
Allele can cause cancer if inserted into wrong place
Inserted allele can get over expressed
Somatic theory effects short lived
DNA sequencing mixture
DNA primers Free nucleotides Modified nucleotide with fluorescent DNA polymerase Single stranded DNA fragments
How is DNA sequenced? ▪ amplify? ▪ what forms? ▪ separation? ▪ identification?
PCR used to amplify DNA fragments
Modified nucleotides cause DNA of different length to be formed (when it joins it causes termination of sequence)
Electrophoresis used to separate fragments
UV light shone on gel forming bands (read bottom up)
How is genome sequenced? ▪ cut ▪ inserted inserted ▪ multiply ▪ extraction ▪ sequenced ▪ analysed
Restriction enzymes cut genome into smaller fragments
Inserted into different BAC which is inserted into bacteria
Left to colonise
DNA extracted using restriction enzyme
Sequenced via chain termination
Combined using computer
Evolutionary relationships
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All organisms share common ancestor
Closely related species share more common DNA
Whole genomes can be sequenced and compared to see how closely related
Comparing genomes of members of same species tells us about evolutionary relationships
