Polymerase Chain Reaction (PCR) in addition to its Applications Amplifies a small, specific region of DNA PCR Applications of PCR

Polymerase Chain Reaction (PCR) in addition to its Applications Amplifies a small, specific region of DNA PCR Applications of PCR www.phwiki.com

Polymerase Chain Reaction (PCR) in addition to its Applications Amplifies a small, specific region of DNA PCR Applications of PCR

Durst, Will, Contributor has reference to this Academic Journal, PHwiki organized this Journal Polymerase Chain Reaction (PCR) in addition to its Applications Lesson Plan 1 PCR What does it do What do Scientists use it as long as The reaction sample in detail How to use micropipettes Practical – Set up your own PCR reactions in addition to amplify them in the Thermal Cycler Gel electrophoresis – How to visualise the DNA that has been amplified in your PCR reactions Lesson Plan 2 Practical – load your amplified PCR samples onto a gel in addition to apply electric current to the gel to separate the DNA fragments according to their size Using PCR to detect faulty BRCA2 genes in addition to how these genes are involved in breast cancer Practical – visualise the DNA fragments that have been separated by gel electrophoresis Interpret the results

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Amplifies a small, specific region of DNA PCR Conservation of endangered species Applications of PCR .CAGTCGCTAAGTTCTAACGTCC PCR: The Steps Find out the sequence of the piece of DNA that you would like to amplify Design & add a short piece of DNA (a PRIMER) which is perfectly complimentary to that sequence

Primers – short single-str in addition to ed pieces of DNA which are chosen to PERFECTLY MATCH a portion of the DNA segment to be copied. There is a FORWARD (F) in addition to a REVERSE (R) primer. DNA F R .CAGTCGCTAAGTTCTAACGTCC G G G A C PCR: The Steps This then allows a whole new complimentary str in addition to to be produced 4. The process can be repeated many times to produce lots of new copies of the original DNA Split the double-str in addition to ed DNA into 2 single str in addition to s (this is called DENATURATION) Join on short pieces of DNA (primers) which perfectly match the denatured DNA (this is called ANNEALING) Extend the DNA to make a perfect copy of the single str in addition to (EXTENSION) PCR

Temperature PCR only works if the temperature is correct in addition to accurate as long as each step Denaturation: 95ºC Annealing: 50 – 65ºC Extension: 72ºC Temperature – Thermal Cycler Accurate temperatures Fast changes In the Reaction Tube Buffer – a chemical that enables the PCR reaction to take place. Has optimal pH in addition to salt components Source of DNA Primers MgCl – Magnesium chloride is a salt which is crucial as long as the DNA polymerase enzyme to work

In the Reaction Tube Nucleotides – free nucleotides (A, T, C & G) are needed to EXTEND the DNA chain on each cycle Taq DNA Polymerase – enzyme used to read the original DNA segment in addition to add on new nucleotides to make a complimentary copy of that sequence Taq DNA Polymerase The Taq (Thermus aquaticus) DNA polymerase has to be a special heat-stable enzyme which is able to survive at high temperature Hot Springs at Yellowstone National Park, USA. Image by Billy Gast. Used under licence. How to avoid the DNA polymerase being broken down in the thermal cycler DNA Polymerase

Taq DNA polymerase is an example of a big improvement Be as long as e its discovery, normal DNA polymerase was used in addition to had to be added freshly after each amplification cycle (since it was degraded at high temperatures) This has made the procedure less labour intensive in addition to cheaper Improvements

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