<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-polymerase-chain-reaction-primary"><primary style="font-size:24px"> Polymerase Chain Reaction </primary></h3> <hr> <main> <ul> <li>Polymerase Chain Reaction (PCR)</li> <li><strong>Definition</strong>: a laboratory technique used to amplify a specific segment of DNA</li> <li>Invented by Kary Mullis in 1983</li> <li>Used in various applications, such as genetic testing, gene cloning, and forensics</li> <li><strong>PCR can be done in three steps</strong>: DNA Denaturation, Annealing, and Extension</li> </ul> </main> <hr> <footer style = "font-size: 18px"> $\rightarrow$ $\rightarrow$ <span style = "color:blue"> Polymerase Chain Reaction </span> $\rightarrow$ PCR step 1 DNA Denaturation $\rightarrow$ PCR step 2 Annealing </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-1"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-step-1-dna-denaturation-primary"><primary style="font-size:24px"> PCR step 1 DNA Denaturation </primary></h3> <hr> <main> <ul> <li>DNA double helix is heated to separate the two strands</li> <li>Denaturation temperature is typically 94-98°C</li> <li>Hydrogen bonds between base pairs are broken</li> <li>Two separate DNA strands are formed</li> </ul> </main> <hr> <footer style = "font-size: 18px"> $\rightarrow$ Polymerase Chain Reaction $\rightarrow$ <span style = "color:blue"> PCR step 1 DNA Denaturation </span> $\rightarrow$ PCR step 2 Annealing $\rightarrow$ PCR step 2 Annealing (Contd.) </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-2"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-step-2-annealing-primary"><primary style="font-size:24px"> PCR step 2 Annealing </primary></h3> <hr> <main> <ul> <li>Temperature is reduced to allow the primers to bind to the DNA template</li> <li>Primers are short DNA sequences that are complementary to the target DNA region</li> <li>They provide a starting point for DNA synthesis</li> <li>Primers bind to the specific DNA sequence of interest</li> </ul> </main> <hr> <footer style = "font-size: 18px">Polymerase Chain Reaction $\rightarrow$ PCR step 1 DNA Denaturation $\rightarrow$ <span style = "color:blue"> PCR step 2 Annealing </span> $\rightarrow$ PCR step 2 Annealing (Contd.) $\rightarrow$ PCR step 3 Extension </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-3"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-step-2-annealing-contd-primary"><primary style="font-size:24px"> PCR step 2 Annealing (Contd.) </primary></h3> <hr> <main> <ul> <li>Annealing temperature is typically 50-65°C</li> <li>Primers attach to the single-stranded DNA template through hydrogen bonding</li> <li>Primers bind to the 3’ end of the target DNA sequence in opposite directions</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR step 1 DNA Denaturation $\rightarrow$ PCR step 2 Annealing $\rightarrow$ <span style = "color:blue"> PCR step 2 Annealing (Contd.) </span> $\rightarrow$ PCR step 3 Extension $\rightarrow$ PCR step 3 Extension (Contd.) </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-4"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-step-3-extension-primary"><primary style="font-size:24px"> PCR step 3 Extension </primary></h3> <hr> <main> <ul> <li>DNA polymerase synthesizes new DNA strands complementary to the template strands</li> <li>Temperature is increased to allow DNA polymerase to work optimally</li> <li>Extension temperature is typically 70-75°C</li> <li>DNA polymerase adds free nucleotides to the 3’ end of the primers</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR step 2 Annealing $\rightarrow$ PCR step 2 Annealing (Contd.) $\rightarrow$ <span style = "color:blue"> PCR step 3 Extension </span> $\rightarrow$ PCR step 3 Extension (Contd.) $\rightarrow$ Components required for PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-5"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-step-3-extension-contd-primary"><primary style="font-size:24px"> PCR step 3 Extension (Contd.) </primary></h3> <hr> <main> <ul> <li>DNA synthesis occurs in the 5’ to 3’ direction</li> <li>DNA polymerase adds nucleotides to the growing DNA strand</li> <li>The DNA strands are replicated exponentially during each cycle of PCR</li> <li>Each cycle doubles the amount of target DNA</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR step 2 Annealing (Contd.) $\rightarrow$ PCR step 3 Extension $\rightarrow$ <span style = "color:blue"> PCR step 3 Extension (Contd.) </span> $\rightarrow$ Components required for PCR $\rightarrow$ Types of PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-6"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-components-required-for-pcr-primary"><primary style="font-size:24px"> Components required for PCR </primary></h3> <hr> <main> <ul> <li><strong>DNA template</strong>: The DNA sequence to be amplified</li> <li><strong>Primers</strong>: Short DNA sequences that bind to the target DNA region</li> <li><strong>DNA polymerase</strong>: Enzyme that synthesizes new DNA strands</li> <li><strong>dNTPs</strong>: Deoxyribonucleotide triphosphates (A, T, C, G) for DNA synthesis</li> <li><strong>Buffer</strong>: Provides optimal reaction conditions for DNA polymerase</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR step 3 Extension $\rightarrow$ PCR step 3 Extension (Contd.) $\rightarrow$ <span style = "color:blue"> Components required for PCR </span> $\rightarrow$ Types of PCR $\rightarrow$ Applications of PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-7"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-types-of-pcr-primary"><primary style="font-size:24px"> Types of PCR </primary></h3> <hr> <main> <ul> <li><strong>Real-time PCR</strong>: Allows monitoring of the PCR process in real-time</li> <li><strong>Reverse transcription PCR (RT-PCR)</strong>: Converts RNA to complementary DNA (cDNA) for amplification</li> <li><strong>Nested PCR</strong>: Two rounds of PCR to enhance specificity and sensitivity</li> <li><strong>Multiplex PCR</strong>: Simultaneous amplification of multiple target DNA sequences</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR step 3 Extension (Contd.) $\rightarrow$ Components required for PCR $\rightarrow$ <span style = "color:blue"> Types of PCR </span> $\rightarrow$ Applications of PCR $\rightarrow$ Advantages of PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-8"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-applications-of-pcr-primary"><primary style="font-size:24px"> Applications of PCR </primary></h3> <hr> <main> <ul> <li><strong>Genetic testing</strong>: Used for identifying genetic disorders or mutations</li> <li><strong>Gene cloning</strong>: Amplifies and produces large amounts of DNA for further studies</li> <li><strong>Forensic analysis</strong>: Amplifies DNA from crime scene samples for identification</li> <li><strong>Diagnosis of infectious diseases</strong>: Detects pathogens in patient samples</li> </ul> </main> <hr> <footer style = "font-size: 18px">Components required for PCR $\rightarrow$ Types of PCR $\rightarrow$ <span style = "color:blue"> Applications of PCR </span> $\rightarrow$ Advantages of PCR $\rightarrow$ PCR Primers </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-9"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-advantages-of-pcr-primary"><primary style="font-size:24px"> Advantages of PCR </primary></h3> <hr> <main> <ul> <li><strong>High sensitivity</strong>: Can detect minute amounts of DNA</li> <li><strong>Rapid and efficient</strong>: Amplification can be completed within hours</li> <li><strong>Specific</strong>: Primers ensure amplification of target DNA sequence only</li> <li><strong>Versatile</strong>: Can be used for a wide range of applications</li> <li><strong>Cost-effective</strong>: Requires minimal equipment and reagents</li> </ul> </main> <hr> <footer style = "font-size: 18px">Types of PCR $\rightarrow$ Applications of PCR $\rightarrow$ <span style = "color:blue"> Advantages of PCR </span> $\rightarrow$ PCR Primers $\rightarrow$ PCR Cycling Parameters </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-10"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-primers-primary"><primary style="font-size:24px"> PCR Primers </primary></h3> <hr> <main> <ul> <li>Short DNA sequences</li> <li>Typically 18-25 nucleotides long</li> <li>Designed to be complimentary to the target DNA sequence</li> <li>Important for specificity of amplification</li> <li>Forward and reverse primers used in PCR</li> </ul> </main> <hr> <footer style = "font-size: 18px">Applications of PCR $\rightarrow$ Advantages of PCR $\rightarrow$ <span style = "color:blue"> PCR Primers </span> $\rightarrow$ PCR Cycling Parameters $\rightarrow$ PCR Optimization </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-11"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-cycling-parameters-primary"><primary style="font-size:24px"> PCR Cycling Parameters </primary></h3> <hr> <main> <ul> <li>Denaturation: 94-98°C for 30 seconds to 2 minutes</li> <li>Annealing: 50-65°C for 30 seconds to 1 minute</li> <li>Extension: 70-75°C for 1-2 minutes</li> <li>Number of cycles: usually 25-35</li> <li>Parameters can be optimized based on the specific target DNA and primer requirements</li> </ul> </main> <hr> <footer style = "font-size: 18px">Advantages of PCR $\rightarrow$ PCR Primers $\rightarrow$ <span style = "color:blue"> PCR Cycling Parameters </span> $\rightarrow$ PCR Optimization $\rightarrow$ Gel Electrophoresis </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-12"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-pcr-optimization-primary"><primary style="font-size:24px"> PCR Optimization </primary></h3> <hr> <main> <ul> <li>Adjusting reaction conditions for optimal results</li> <li>Factors to consider: <ul> <li>Primer concentration</li> <li>Mg2+ concentration</li> <li>Annealing temperature</li> <li>Extension time</li> <li>DNA template quality and concentration</li> </ul> </li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR Primers $\rightarrow$ PCR Cycling Parameters $\rightarrow$ <span style = "color:blue"> PCR Optimization </span> $\rightarrow$ Gel Electrophoresis $\rightarrow$ Gel Electrophoresis (Contd.) </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-13"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-gel-electrophoresis-primary"><primary style="font-size:24px"> Gel Electrophoresis </primary></h3> <hr> <main> <ul> <li>Technique used to separate DNA fragments by size</li> <li>DNA samples are loaded onto an agarose gel</li> <li>Applied electric current causes DNA to move towards the positive electrode</li> <li>Smaller DNA fragments move faster and travel further than larger fragments</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR Cycling Parameters $\rightarrow$ PCR Optimization $\rightarrow$ <span style = "color:blue"> Gel Electrophoresis </span> $\rightarrow$ Gel Electrophoresis (Contd.) $\rightarrow$ Quantitative PCR (qPCR) </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-14"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-gel-electrophoresis-contd-primary"><primary style="font-size:24px"> Gel Electrophoresis (Contd.) </primary></h3> <hr> <main> <ul> <li>After electrophoresis, DNA fragments can be visualized using a DNA stain (e.g., ethidium bromide)</li> <li>DNA bands can be compared to size markers to determine fragment size</li> <li>Gel electrophoresis is commonly used to analyze PCR products</li> </ul> </main> <hr> <footer style = "font-size: 18px">PCR Optimization $\rightarrow$ Gel Electrophoresis $\rightarrow$ <span style = "color:blue"> Gel Electrophoresis (Contd.) </span> $\rightarrow$ Quantitative PCR (qPCR) $\rightarrow$ Reverse Transcription PCR (RT-PCR) </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-15"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-quantitative-pcr-qpcr-primary"><primary style="font-size:24px"> Quantitative PCR (qPCR) </primary></h3> <hr> <main> <ul> <li>Also known as real-time PCR</li> <li>Monitors the amplification of DNA in real-time</li> <li>Uses a fluorescent probe that emits fluorescence when bound to the amplified DNA</li> <li>Measures the number of PCR cycles required for the fluorescence to reach a certain threshold</li> </ul> </main> <hr> <footer style = "font-size: 18px">Gel Electrophoresis $\rightarrow$ Gel Electrophoresis (Contd.) $\rightarrow$ <span style = "color:blue"> Quantitative PCR (qPCR) </span> $\rightarrow$ Reverse Transcription PCR (RT-PCR) $\rightarrow$ Nested PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-16"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-reverse-transcription-pcr-rt-pcr-primary"><primary style="font-size:24px"> Reverse Transcription PCR (RT-PCR) </primary></h3> <hr> <main> <ul> <li>Converts RNA to complementary DNA (cDNA) for amplification</li> <li>Utilizes the enzyme reverse transcriptase to synthesize cDNA</li> <li>Allows for the detection and quantification of RNA transcripts</li> <li>Widely used for gene expression analysis</li> </ul> </main> <hr> <footer style = "font-size: 18px">Gel Electrophoresis (Contd.) $\rightarrow$ Quantitative PCR (qPCR) $\rightarrow$ <span style = "color:blue"> Reverse Transcription PCR (RT-PCR) </span> $\rightarrow$ Nested PCR $\rightarrow$ Multiplex PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-17"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-nested-pcr-primary"><primary style="font-size:24px"> Nested PCR </primary></h3> <hr> <main> <ul> <li>Two rounds of PCR amplification</li> <li>First round uses outer primers to amplify a larger DNA fragment</li> <li>Second round uses inner primers that are complementary to the first round PCR product</li> <li>Increases specificity and sensitivity</li> <li>Useful when amplifying low abundant DNA targets</li> </ul> </main> <hr> <footer style = "font-size: 18px">Quantitative PCR (qPCR) $\rightarrow$ Reverse Transcription PCR (RT-PCR) $\rightarrow$ <span style = "color:blue"> Nested PCR </span> $\rightarrow$ Multiplex PCR $\rightarrow$ Applications of PCR </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-18"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-multiplex-pcr-primary"><primary style="font-size:24px"> Multiplex PCR </primary></h3> <hr> <main> <ul> <li>Simultaneous amplification of multiple DNA targets in a single reaction</li> <li>Uses different sets of primers, each specific to a target DNA sequence</li> <li>Amplified products can be distinguished based on size or specific fluorescent probes</li> <li>Saves time and resources by reducing the number of reactions required</li> </ul> </main> <hr> <footer style = "font-size: 18px">Reverse Transcription PCR (RT-PCR) $\rightarrow$ Nested PCR $\rightarrow$ <span style = "color:blue"> Multiplex PCR </span> $\rightarrow$ Applications of PCR $\rightarrow$ </footer>

<h3 id="success-stylefont-size25px-biotechnology-principles-and-processes-polymerase-chain-reaction-success-19"><Success style="font-size:25px"> Biotechnology Principles And Processes Polymerase Chain Reaction </Success></h3> <h3 id="primary-stylefont-size24px-applications-of-pcr-primary-1"><primary style="font-size:24px"> Applications of PCR </primary></h3> <hr> <main> <ul> <li>Medical diagnostics: identification of pathogens, genetic disorders, and cancer markers</li> <li>Forensic analysis: DNA profiling for crime scene investigations</li> <li>Agricultural biotechnology: GMO detection and crop improvement</li> <li>Environmental research: assessing biodiversity and monitoring species abundance</li> <li>Evolutionary biology: studying DNA to understand evolutionary relationships</li> </ul> </main> <hr> <footer style = "font-size: 18px">Nested PCR $\rightarrow$ Multiplex PCR $\rightarrow$ <span style = "color:blue"> Applications of PCR </span> $\rightarrow$ $\rightarrow$ </footer>