6.1.3 - Manipulating Genomes Flashcards
What is a genome?
all of the DNA in an organism
- from nucleus and mitochondria
- human genome has 3.5 million base pairs in DNA
What is DNA sequencing?
working out the sequence of bases in a strand of DNA
What new developments have increased the speed of DNA sequencing/processing?
- high throughput sequencing
- shotgun sequencing
- whole genome sequencing
- next generation sequencing
- pyrosequencing (using luciferase)
- massive parallel sequencing
Why is gene sequencing important?
- allows for genome-wide comparison between individuals and species
- allows for sequences of amino acids in polypeptides to be predicted
- allows for development of synthetic biology
What is bioinformatics?
the development of the software and computing skills needed to analyse and organise raw biological data
eg. development of mathematical models, statistical tests, algorithms, etc.
What is computational biology?
the study of biology using computational techniques to analyse large amounts of data
-uses data to build theoretical models of biological systems, which can be used to predict outcomes in different situations
What is genomics?
the field of genetics which applies DNA sequencing methods and computational biology to analyse the structure and function of genomes
How has gene sequencing allowed for genome-wide comparisons between individuals and between species?
- computers can analyse and compare genomes
- patterns in DNA inherited can be revealed
- patterns in diseases we are vulnerable to can be revealed (aids medicine and epidemiology research)
- aids identification of species (by comparison to a standard, eg. in DNA barcoding)
- evolutionary relationships between organisms can be understood better
What is synthetic biology?
the design and construction of biological pathways, organisms or devises, or the redesign of existing natural biological systems
What techniques are considered a part of synthetic biology?
- genetic engineering
- synthesis of new genes to replace faulty genes
- synthesis of a new organism
- use of biological systems in industrial contexts (like using immobilised enzymes)
What is DNA profiling?
producing an image of the patterns in the non-coding DNA of an individual
-uses mini and micro satellites instead of genes
How is a DNA profile produced?
- DNA is extracted (using a tissue sample)
- sample is digested (broken down using restriction endonucleases to cut the DNA at specific points along the nucleotide sequence, known as restriction sites)
- DNA fragments are separated (using electrophoresis)
- hybridisation (DNA probes with radioactive or fluorescent markers are used to highlight important DNA sequences)
- evidence is viewed (using images from DNA probes or sequencing data)
What are the uses of DNA profiling?
- forensics
- analysis of disease risk
- paternity tests
What are restriction endonucleases?
enzymes which cut DNA at specific points (restriction sites) into smaller pieces
- different enzymes cut at different sites
- used in DNA profiling
What does PCR stand for?
polymerase chain reaction
What is the polymerase chain reaction (PCR)?
a process which amplifies a small sample of DNA
- produces a large number of samples of the same section of DNA, which can then be used in further study
- uses specific enzymes and temp changes
What happens in the polymerase chain reaction (PCR)?
- strands are separated (temp is increased to 95°C to denature the DNA by breaking the H-bonds between the complementary bases to separate the 2 strands)
- annealing of primers (temp is decreased to 55°C and primers anneal/bind to the end of the sections of DNA that you want to amplify)
- DNA synthesis (temp is increased to 72°C and free DNA nucleotides complementary base pair to exposed bases and Taq polymerase adds these bases onto the primer by forming phosphodiester bonds to produce a double-stranded sample of DNA identical to the desired sequence)
- this is repeated to produce lots of samples of the desired sequence
How are the strands of DNA separated in PCR?
- temp is increased to 95°C
- DNA is denatures as the H-bonds between the complementary bases are broken
How do primers anneal to DNA in PCR?
- temp is decreased to 55°C
- primers anneal/bind to the end of the sections of DNA that you want to amplify by complementary base pairing
How is DNA synthesised in PCR?
- temp is increased to 72°C
- free DNA nucleotides complementary base pair to exposed bases
- Taq polymerase adds these bases onto the primer by forming phosphodiester bonds to produce a double-stranded sample of DNA identical to the desired sequence
What temperature is required during PCR for the DNA strands to be separated?
95°C
What temperature is required during PCR for the annealing of primers?
55°C
What temperature is required during PCR for the synthesis of DNA?
72°C
Why is Taq polymerase used in PCR rather than DNA polymerase?
Taq polymerase is extracted from thermophilic bacteria found in hot springs so is adapted to high temperatures (so won’t denature)
