Applications of genetics Flashcards
What is Sanger sequencing?
Using chemically altered bases which cause DNA polymerase to stop when base is incorporated
What are the aims of the human genome project?
-Determine sequences of 3 billion base pairs in human DNA
-Identify all gene loci, and which chromosome it’s on
-Understand coding for amino acids and protein
-Store info in databases
What are the uses of the human genome project?
-Better prediction of the effect of drugs
-Improved design of drugs
-Development of new and improved treatment of disease
What are the limitations of the human genome project?
-Sample size too small
-Doesn’t represent different ethnicities
-Anonymous, so we don’t know the health of individuals
What are the aims of the 100k genome project?
-Study variation
-Locate genes responsible for conditions
-Research treatments
What are the strengths of the 100k genome project?
-Sample of people selected based on cancer/disorder status
-Larger sample to identify variation
-More ages, sex, ethnicities
What are the limitations of the 100k genome project?
-Doesn’t include Africa and S. America
-Not representative of human population
What are the ethical issues of genome sequencing?
1) concern of ownership of data
2) Cherry picking embryos
3) Screening can cause anxiety
4) Concerns over storage
Why was Anopheles gambiae sequenced?
-Rapid evolution of insecticide resistance
-Develop new chemicals to kill mosquito and lower transmission
Why was plasmodium sequenced?
-Developing multidrug resistance
-Develop more effective drugs for humans
What goes into the PCR machine?
-Primers
-Nucleotides
-Gene
-TAQ polymerase
-Buffer
What is the method for PCR?
1) 95c: Separates DNA into 2 strands
2) 55c: Allow primers to anneal to complimentary bases and start replication
3) DNA polmerase synthesizes complimentary strand, forming phosphodiester bonds
What are the limitations of PCR?
-Contamination
-Error rate
-Limited amplification
-Suitable DNA fragment size
-Sensitivity to inhibitors
What is the method for gel electrophoresis?
1) DNA is extracted using restriction endonuclease, and loaded into wells
2) Gel exposed to electric current
3) Negatively charged DNA fragments move towards positive terminal
4) Smaller fragments move longer distance in same time
5) Use DNA ladder to estimate fragment size
What can be used to visualize gel electrophoresis?
-Blue dye
-Radioactive probes
What can DNA profiling be used for?
-Paternity testing
-Identify dead bodies
-Store genetic info
-Composition of food
What is the definition of genetic engineering?
Transfer of a gene from one organism to another gene expressed in a new host cell (transgenic/recombinant)
What are the steps of genetic engineering?
1) Identify and isolate gene
2) PCR
3) Insert gene into vector
4) Insert Vector into host cell
5) Production of protein by host cell (transcribe and translate)
Describe the use of restriction endonuclease
-A bacterial enzyme that will cut DNA at a specific nucleotide sequence
-Cut phosphodiester bonds staggered, creating sticky ends
What are the limitations of restriction endonuclease?
-Need to know gene loci
-Could cut within gene
-Eukaryotic genes contain introns, whilst prokaryotes don’t, so bacteria may not have the enzymes needed to processs pre mRNA
-Any proteins with AA aren’t functional
Describe the use of Reverse transcriptase
-Extract mRNA
-Reverse transcriptase makes complimentary single stranded cDNA
-DNA polymerase makes complimentary double stranded DNA
-Sticky ends added
Describe the use of viral vectors
-Infect host cell with genetic information
-Concerns over pathogen and fast mutation rate
Describe the use of plasmids
1) Bacteria cell wall and membrane destabilised
2) Plasmids isolated from bacteria
3) Plasmid cut open using same RE (same stickey ends)
4) DNA ligase joins gene to plasmid, forming phosphodiester bonds
Describe the purification of recombinant genes
-Bacteria cells put into fermenter
-Transcribe and translate
