Polymerase Chain Reaction (PCR) | Cheat Sheet - Edubirdie

University:
University of California, San Diego
Course:
BIMM 100 | Molecular Biology
Academic year:

2024
Views:

206

Pages:

2
Author:

customer-8930767

principles and applications of the polymerase chain reaction (PCR), including thermal cycling parameters and detection methods. Polymerase chain reaction (PCR) is an in vitro DNA amplification method using thermal cycling and a thermostable DNA polymerase (e.g., Taq from Thermus aquaticus). It exponentially amplifies a specific target sequence defined by two oligonucleotide primers. Components: DNA template, forward/reverse primers (18–25 nt), dNTPs, Mg²? (cofactor), buffer, DNA polymerase. Thermal cycling (typical 30–40 cycles): Denaturation (94–98°C ): Breaks H-bonds, yielding single-stranded DNA. Annealing (50–65°C ): Primers hybridize to complementary sequences. Annealing temperature (T?) calculated by base composition (e.g., 2°C × (A+T) + 4°C × (G+C)). Extension (72°C ): Polymerase adds dNTPs from primers, synthesizing new strands. Taq has 5’?3’ polymerase but lacks proofreading (error ~1/10? bases). High-fidelity polymerases (e.g., Pfu) have 3’?5’ exonuclease proofreading. Theoretical amplification: 2? copies after n cycles (e.g., 2³? ? 1 billion copies). Plateau occurs due to reagent depletion and enzyme inactivation. Detection methods: End-point: Gel electrophoresis with intercalating dye (ethidium bromide, SYBR Green) or silver staining. Real-time PCR (qPCR ): Monitors amplification in real time using fluorescent probes (TaqMan—hydrolysis probe with quencher) or SYBR Green (binds dsDNA). Quantifies starting template via Ct (threshold cycle) value, using standard curve or ??Ct method. Applications: DNA fingerprinting, pathogen detection (HIV, SARS-CoV-2), mutation analysis (allelespecific PCR), sequencing template preparation, ancient DNA amplification, and RT-PCR (reverse transcriptase PCR) for RNA expression. Limitations: Requires known target sequences; contamination risk; inhibitor sensitivity (heme, phenol); amplicon length typically <5 kb.

Related Documents

Polymerase Chain Reaction (PCR)

Polymerase Chain Reaction (PCR) - Page 1

Polymerase Chain Reaction (PCR) - Page 2

of 2

Recommended Documents

Chapter 18: The Endocrine System

Chapter 18: The Endocrine System

University of California, San Diego BIMM 100 | Molecular Biology

Study Guide

Cellular Respiration Virtual Lab Answer Key

Cellular Respiration Virtual Lab Answer Key

University of California, San Diego BIMM 100 | Molecular Biology

Answer Key

Biology Culminating Tasks

Biology Culminating Tasks

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Natural Selection Academic Notes

Natural Selection Academic Notes

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

AP Biology 2022 Free-Response Questions

AP Biology 2022 Free-Response Questions

University of California, San Diego BIMM 100 | Molecular Biology

Study Guide

Pedigree Analysis Problems

Pedigree Analysis Problems

University of California, San Diego BIMM 100 | Molecular Biology

Answer Key

Symptomatic Cholelithiasis With Suspected Choledocholithiasis

Symptomatic Cholelithiasis With Suspected Choledocholithiasis

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Neurology #2 Long Exam 4

Neurology #2 Long Exam 4

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Benign Gynecology Lesions

Benign Gynecology Lesions

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Vaginal Infections to TSS

Vaginal Infections to TSS

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Cheat Sheet - Clin Path Lecture 1

Cheat Sheet - Clin Path Lecture 1

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Genital Ulcers - 40-item High-yield MCQ Quiz

Genital Ulcers - 40-item High-yield MCQ Quiz

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

New Documents from this Course

Tcell Maturation Lecture Notes

Tcell Maturation Lecture Notes

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Transposable Elements

Transposable Elements

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Retroviral-like Elements

Retroviral-like Elements

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Cytoplasmic Content

Cytoplasmic Content

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Secondary System in Cells

Secondary System in Cells

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Muscle Relaxation Study Handouts

Muscle Relaxation Study Handouts

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Gene Surveillance Mechanism

Gene Surveillance Mechanism

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Co-operativity of Haemoglobin

Co-operativity of Haemoglobin

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Brassinosteroid Study Reference

Brassinosteroid Study Reference

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Plan Response in Hypoxic Conditions

Plan Response in Hypoxic Conditions

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Meristematic Tissue in Plants

Meristematic Tissue in Plants

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note

Signaling Pathways Tutorial Materials

Signaling Pathways Tutorial Materials

University of California, San Diego BIMM 100 | Molecular Biology

Lecture Note