heat-tolerant bacterium thesis
heat-tolerant bacterium thesis
Methods and the Human Genome Project
What is PCR?
- Polymerase chain reaction (PCR) is a common lab technique used to make millions or billions of copies of a particular region of DNA
- This region can be anything the experimenter wants to know more about – it could be a gene with a function a researcher wants to understand
- It is an in vitro practice
- PCR requires a DNA polymerase enzyme that makes new strands of DNA, using the existing strands as templates – just like in DNA replication in an organism. In PCR, this DNA polymerase is usually Taq polymerase
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- Taq is named after the heat-tolerant bacterium it was isolated from (thermus aquaticus)
- Interestingly, Taq was first discovered at Yellowstone National Park
- Thermus aquaticus is a thermophile bacterium. is stable at elevated temperatures.
- DNA using Taq is achieved at the optimal temperature of 70°C and is used during PCR to denature the template DNA, or separate its strands
- Taq polymerase can only make DNA if it’s given a primer – a short sequence of nucleotides that provides a starting point for DNA synthesis
- PCR amplification is achieved by using oligonucleotide primers for flanking regions with a known base sequence.
- PCR primers are short pieces of single-stranded DNA, usually around 20 nucleotides in length
- Two primers are used per each PCR reaction. They flank the target region to be copied – aka, they are given sequences that cause them to bind to opposite strands of the template DNA.
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Image source: Khan Academy
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- Taq polymerase can only make DNA if it’s given a primer – a short sequence of nucleotides that provides a starting point for DNA synthesis
- PCR amplification is achieved by using oligonucleotide primers for flanking regions with a known base sequence.
- PCR primers are short pieces of single-stranded DNA, usually around 20 nucleotides in length
- Two primers are used per each PCR reaction. They flank the target region to be copied – aka, they are given sequences that cause them to bind to opposite strands of the template DNA.
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- Major components
- Taq polymerase
- Primers
- Template DNA
- Nucleotides
- Assemble these components in a tube and put through repeated cycles of heating/cooling to allow for DNA synthesis to occur
- Basic steps:
Denaturation: Heating the reaction will separate (denature) the DNA strands to provide the single-stranded template for step 2.
Annealing: Cooling the reaction so primers can bind to their complementary sequences on the single-stranded template DNA.
