6.1.3 Flashcards
What is the Sanger chain termination technique?
1) extract DNA and cut it into fragments of various lengths. Amplify
2) Sequence the DNA by adding it to 4 different solutions, DNA nucleotides, DNA polymerase, primers and a terminator base
3) Electrophoresis
What do you do after extracting the DNA?
cut into fragments of various lengths
What do you do after cutting DNA into fragments?
amplify
What happens after amplifying DNA fragments?
sequence DNA by adding it to 4 different solutions
What do the solutions in Sanger technique contain?
terminator base
DNA nucleotides
DNA polymerase
Primers
What happens after sequencing the DNA?
electrophoresis
What will electrophoresis do?
separates DNA depending on mass
What is a faster technique for DNA sequencing?
Massive parallel sequencing
Next gen sequencing
High throughout sequencing
What do we do after sequencing?
take solutions that have our bases in and place them into wells in electrophoresis
Which direction should the current be in for electrophoresis?
negative to positive
How do DNA fragments move through electrophoresis?
DNA is slightly negatively charged so repelled by cathode and attracted to anode
Why do smaller fragments travel futher up?
less resistance
What are 2 ways to see DNA?
Southern blotting using radioactive DNA probes and X rays
Using green fluorescent protein , DNA probe and UV light
What is bioinformatics?
software is developed to process and understand large complex data using computational biology
What does bioinformatics allow?
acsess to large amounts of data
What is compuational biology?
acsess to large amounts of data on DNA and proteins
information is universal
allows rapid comparison of sequencs with newly sequenced alleles
amino acid sequence / protein structures held in database
computer modelling of new protein structure from base sequence
What does bioinformatics allow the rapid comparison of?
sequences and newly-sequenced allels
What is synthetic biology?
field of science
involves redesigning organisms for useful purposes by engineering them to have new abilities
to solve problems in medicine, manufactiring and medicine
What can synthetic biology do?
problems in medicine
manufacturing and agriculture
What is bioinformatic used in?
epidemology
How can bioinformatics be used for in epidemology?
identfiy source o outbreak
identfiy bulnerable pop
esign vaccination programmes t target certain individuald
What is proteomic?
large scale study of a set f proteins in an organisms
Why do we compare genomes?
universal
look at phylogeny
What are the uses of DNA profiling?
paternity testing
Why are introns used in DNA profiling?
in most people the genome is very similar
using coding sequences of DNA would not proide unique profiles
non-coding DNA contains VNTR / STR / repeating sequences
What are the stages of DNA profiling?
Amplify DNA using PCR
Cut DNA using RE at specific areas
Put DNA onto gel electrophoresis
separate DNA based on mass
visualise using UV light
Outline genetic engineering?
Restriction endonucleases are used to cut desired gene from DNA
What do restriction endonucleases do?
cut the desired gene from DNA
What does restriction endonucleases cutting desired genes from DNA cause?
Sticky ends to be created
What does sticky ends allow?
makes it easier to insert desired gene
What is the plasmid / vector cut with?
same restriction enzymes to produce complementary sticky ends
What does the DNA ligase do?
helps insert desired gene into the plasmid
What is added to a desired gene when it is inserted into the plasmid?
marker
What does the marker do?
fluorescent marker to see if desired gene has been taken up by vector
What type of DNA is inserted into the host cell?
recombinantq
What is electroporation?
electric shock makes membrane porous so plasmids can pass through membrane of the host cell
What process does the host cell undergo?
mitosis
What is recombinant DNA?
DNA combined from 2 sources
What are the vectors in plants, animals and bacteria?
Virus
Outline genetic engineering?
1) Restriction endonucleases cut desired gene from DNA
2) creates sticky ends which make it easier to insert desired genes
3)plasmid cut using same restriction enzymes to produce complementary sticky ends
4)desired gene inserted into plasmid / vector using DNA ligase. desired gene inserted with a marker
5) recombinant DNA inserted into host cell using electroporation
6) host cell undergoes mitosis reproducing desired gene
Why do we use PCR?
Amplify DNA
Outline PCR?
Denaturation
heat 94-98 degrees
breaks H2 bonds
Annealing
cool 55-70 degrees
primer needed to allow Taq polymerase to join
Extension
66-72 degrees
heated to the optimum temp for the enzyme
Where is Taq polymerase found?
extremophiles
bacterial that live in hot springs
