Molecular Diagnostics

  1. Core Methodologies and Technologies
    1. Sample Collection and Processing
      1. Sample Types and Requirements
        1. Blood Specimens
          1. Whole Blood
            1. Plasma
              1. Serum
                1. Buffy Coat
                  1. Collection Tubes and Anticoagulants
                  2. Tissue Specimens
                    1. Fresh Tissue
                      1. Frozen Tissue
                        1. Formalin-Fixed Paraffin-Embedded Tissue
                          1. Tissue Processing Considerations
                          2. Body Fluids
                            1. Cerebrospinal Fluid
                              1. Urine
                                1. Saliva
                                  1. Amniotic Fluid
                                    1. Pleural Fluid
                                    2. Non-Invasive Samples
                                      1. Buccal Swabs
                                        1. Stool Samples
                                          1. Cell-Free DNA Sources
                                        2. Pre-Analytical Considerations
                                          1. Sample Stability
                                            1. Storage Conditions
                                              1. Transport Requirements
                                                1. Chain of Custody
                                                2. Nucleic Acid Extraction
                                                  1. DNA Extraction Methods
                                                    1. Organic Extraction
                                                      1. Silica-Based Purification
                                                        1. Magnetic Bead Separation
                                                          1. Automated Extraction Systems
                                                          2. RNA Extraction Methods
                                                            1. Phenol-Based Extraction
                                                              1. Column-Based Purification
                                                                1. RNA Stabilization
                                                                  1. RNase Contamination Prevention
                                                                  2. Quality Assessment
                                                                    1. Spectrophotometric Analysis
                                                                      1. Fluorometric Quantification
                                                                        1. Gel Electrophoresis
                                                                          1. Bioanalyzer Analysis
                                                                      2. Polymerase Chain Reaction Technologies
                                                                        1. Basic PCR Principles
                                                                          1. Thermal Cycling
                                                                            1. Denaturation
                                                                              1. Annealing
                                                                                1. Extension
                                                                                2. Essential Components
                                                                                  1. Template DNA
                                                                                    1. Primers
                                                                                      1. DNA Polymerase
                                                                                        1. Deoxynucleotides
                                                                                          1. Buffer Systems
                                                                                            1. Magnesium Chloride
                                                                                          2. PCR Optimization
                                                                                            1. Primer Design
                                                                                              1. Specificity Considerations
                                                                                                1. Melting Temperature Calculations
                                                                                                  1. Primer-Dimer Avoidance
                                                                                                  2. Reaction Conditions
                                                                                                    1. Temperature Profiles
                                                                                                      1. Cycle Numbers
                                                                                                        1. Reagent Concentrations
                                                                                                      2. PCR Variants
                                                                                                        1. Real-Time PCR
                                                                                                          1. Fluorescent Detection Systems
                                                                                                            1. Quantification Methods
                                                                                                              1. Melting Curve Analysis
                                                                                                              2. Reverse Transcription PCR
                                                                                                                1. cDNA Synthesis
                                                                                                                  1. One-Step vs Two-Step Protocols
                                                                                                                    1. RNA Template Considerations
                                                                                                                    2. Multiplex PCR
                                                                                                                      1. Multiple Target Amplification
                                                                                                                        1. Primer Compatibility
                                                                                                                          1. Detection Strategies
                                                                                                                          2. Nested PCR
                                                                                                                            1. Two-Round Amplification
                                                                                                                              1. Contamination Prevention
                                                                                                                              2. Digital PCR
                                                                                                                                1. Reaction Partitioning
                                                                                                                                  1. Absolute Quantification
                                                                                                                                    1. Droplet Digital PCR
                                                                                                                                  2. Isothermal Amplification
                                                                                                                                    1. Loop-Mediated Isothermal Amplification
                                                                                                                                      1. Primer Design Requirements
                                                                                                                                        1. Detection Methods
                                                                                                                                        2. Strand Displacement Amplification
                                                                                                                                          1. Nucleic Acid Sequence-Based Amplification
                                                                                                                                        3. Hybridization-Based Methods
                                                                                                                                          1. Probe Technologies
                                                                                                                                            1. Oligonucleotide Probes
                                                                                                                                              1. DNA Probes
                                                                                                                                                1. RNA Probes
                                                                                                                                                  1. Labeling Methods
                                                                                                                                                    1. Radioactive Labels
                                                                                                                                                      1. Fluorescent Labels
                                                                                                                                                        1. Chemiluminescent Labels
                                                                                                                                                          1. Enzymatic Labels
                                                                                                                                                        2. Blotting Techniques
                                                                                                                                                          1. Southern Blotting
                                                                                                                                                            1. DNA Transfer Methods
                                                                                                                                                              1. Hybridization Conditions
                                                                                                                                                                1. Detection Systems
                                                                                                                                                                2. Northern Blotting
                                                                                                                                                                  1. RNA Transfer Methods
                                                                                                                                                                    1. Probe Hybridization
                                                                                                                                                                      1. Signal Detection
                                                                                                                                                                    2. In Situ Hybridization
                                                                                                                                                                      1. Fluorescence In Situ Hybridization
                                                                                                                                                                        1. Probe Preparation
                                                                                                                                                                          1. Sample Preparation
                                                                                                                                                                            1. Hybridization Protocols
                                                                                                                                                                              1. Signal Detection and Analysis
                                                                                                                                                                              2. Chromogenic In Situ Hybridization
                                                                                                                                                                              3. Microarray Technologies
                                                                                                                                                                                1. DNA Microarrays
                                                                                                                                                                                  1. Spotted Arrays
                                                                                                                                                                                    1. Oligonucleotide Arrays
                                                                                                                                                                                      1. Gene Expression Profiling
                                                                                                                                                                                      2. Comparative Genomic Hybridization
                                                                                                                                                                                        1. Array CGH Principles
                                                                                                                                                                                          1. Copy Number Analysis
                                                                                                                                                                                            1. Data Interpretation
                                                                                                                                                                                        2. DNA Sequencing Technologies
                                                                                                                                                                                          1. First-Generation Sequencing
                                                                                                                                                                                            1. Sanger Sequencing
                                                                                                                                                                                              1. Chain Termination Method
                                                                                                                                                                                                1. Capillary Electrophoresis
                                                                                                                                                                                                  1. Base Calling
                                                                                                                                                                                                    1. Quality Assessment
                                                                                                                                                                                                  2. Next-Generation Sequencing
                                                                                                                                                                                                    1. Platform Technologies
                                                                                                                                                                                                      1. Illumina Sequencing
                                                                                                                                                                                                        1. Ion Torrent Sequencing
                                                                                                                                                                                                          1. 454 Pyrosequencing
                                                                                                                                                                                                          2. Library Preparation
                                                                                                                                                                                                            1. DNA Fragmentation
                                                                                                                                                                                                              1. Adapter Ligation
                                                                                                                                                                                                                1. Size Selection
                                                                                                                                                                                                                  1. Amplification Methods
                                                                                                                                                                                                                  2. Sequencing Strategies
                                                                                                                                                                                                                    1. Whole Genome Sequencing
                                                                                                                                                                                                                      1. Whole Exome Sequencing
                                                                                                                                                                                                                        1. Targeted Panel Sequencing
                                                                                                                                                                                                                          1. RNA Sequencing
                                                                                                                                                                                                                          2. Data Output and Quality
                                                                                                                                                                                                                            1. Read Length and Quality
                                                                                                                                                                                                                              1. Coverage Depth
                                                                                                                                                                                                                                1. Error Rates
                                                                                                                                                                                                                              2. Third-Generation Sequencing
                                                                                                                                                                                                                                1. Single-Molecule Real-Time Sequencing
                                                                                                                                                                                                                                  1. PacBio Technology
                                                                                                                                                                                                                                    1. Long Read Advantages
                                                                                                                                                                                                                                    2. Nanopore Sequencing
                                                                                                                                                                                                                                      1. Oxford Nanopore Technology
                                                                                                                                                                                                                                        1. Real-Time Analysis
                                                                                                                                                                                                                                          1. Portable Sequencing
                                                                                                                                                                                                                                      2. Post-Amplification Analysis
                                                                                                                                                                                                                                        1. Electrophoretic Methods
                                                                                                                                                                                                                                          1. Agarose Gel Electrophoresis
                                                                                                                                                                                                                                            1. Gel Preparation
                                                                                                                                                                                                                                              1. Sample Loading
                                                                                                                                                                                                                                                1. Visualization Methods
                                                                                                                                                                                                                                                2. Polyacrylamide Gel Electrophoresis
                                                                                                                                                                                                                                                  1. High-Resolution Analysis
                                                                                                                                                                                                                                                    1. Denaturing Conditions
                                                                                                                                                                                                                                                    2. Capillary Electrophoresis
                                                                                                                                                                                                                                                      1. Automated Fragment Analysis
                                                                                                                                                                                                                                                        1. Size Determination
                                                                                                                                                                                                                                                          1. Quantitative Analysis
                                                                                                                                                                                                                                                        2. Melting Curve Analysis
                                                                                                                                                                                                                                                          1. High-Resolution Melting
                                                                                                                                                                                                                                                            1. Mutation Scanning
                                                                                                                                                                                                                                                              1. Genotyping Applications
                                                                                                                                                                                                                                                              2. Melting Temperature Determination
                                                                                                                                                                                                                                                              3. Restriction Analysis
                                                                                                                                                                                                                                                                1. Restriction Fragment Length Polymorphism
                                                                                                                                                                                                                                                                  1. Enzyme Selection
                                                                                                                                                                                                                                                                    1. Digestion Conditions
                                                                                                                                                                                                                                                                      1. Fragment Analysis

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                                                                                                                                                                                                                                                                3. Bioinformatics and Data Analysis