Calculations for Molecular Biology and Biotechnology

Chapter 1. Scientific Notation and Metric Prefixes

• Introduction
• 1.1. Significant Digits
• 1.2. Exponents and Scientific Notation
• 1.3. Metric Prefixes
• Chapter Summary

Chapter 2. Solutions, Mixtures, and Media

• Introduction
• 2.1. Calculating Dilutions: A General Approach
• 2.2. Concentrations by a Factor of X
• 2.3. Preparing Percent Solutions
• 2.4. Diluting Percent Solutions
• 2.5. Moles and Molecular Weight—Definitions
• 2.6. Normality
• 2.7. pH
• 2.8. pKa and The Henderson–Hasselbalch Equation
• Chapter Summary

Chapter 3. Cell Growth

• 3.1. The Bacterial Growth Curve
• 3.2. Manipulating Cell Concentration
• 3.3. Plotting OD550 Versus Time on a Linear Graph
• 3.4. Plotting the Logarithm of OD550 Versus Time on a Linear Graph
• 3.5. Plotting the Log of Cell Concentration Versus Time
• 3.6. Calculating Generation Time
• 3.7. Plotting Cell Growth Data on a Semilog Graph
• 3.8. Plotting Cell Concentration Versus Time on a Semilog Graph
• 3.9. Determining Generation Time Directly From a Semilog Plot of Cell Concentration Versus Time
• 3.10. Plotting Cell Density Versus OD550 on a Semilog Graph
• 3.11. The Fluctuation Test
• 3.12. Measuring Mutation Rate
• 3.13. Measuring Cell Concentration on a Hemocytometer
• Chapter Summary

Chapter 4. Working with Bacteriophage

• Introduction
• 4.1. Multiplicity of Infection
• 4.2. Probabilities and Multiplicity of Infection
• 4.3. Measuring Phage Titer
• 4.4. Diluting Bacteriophage
• 4.5. Measuring Burst Size
• Chapter Summary

Chapter 5. Nucleic Acid Quantification

• 5.1. Quantification of Nucleic Acids by Ultraviolet Spectroscopy
• 5.2. Determining the Concentration of Double-Stranded DNA
• 5.3. Determining the Concentration of Single-Stranded DNA Molecules
• 5.4. Oligonucleotide Quantification
• 5.5. Measuring RNA Concentration
• 5.6. Molecular Weight, Molarity, and Nucleic Acid Length
• 5.7. Estimating DNA Concentration on an Ethidium Bromide-Stained Gel
• 5.8. Dye-Labeled Nucleic Acids
• 5.9. Limit of Detection and Limit of Quantitation
• Chapter Summary

Chapter 6. Labeling Nucleic Acids With Radioisotopes

• Introduction
• 6.1. Units of Radioactivity: The Curie
• 6.2. Estimating Plasmid Copy Number
• 6.3. Labeling DNA by Nick Translation
• 6.4. Random Primer Labeling of DNA
• 6.5. Labeling 3′ Termini With Terminal Transferase
• 6.6. Complementary DNA Synthesis
• 6.7. Homopolymeric Tailing
• 6.8. In Vitro Transcription
• Chapter Summary

Chapter 7. Oligonucleotide Synthesis

• Introduction
• 7.1. Synthesis Yield
• 7.2. Measuring Stepwise and Overall Yield by the Dimethoxytrityl Cation Assay
• 7.3. Calculating Micromoles of Nucleoside Added at Each Base Addition Step
• Chapter Summary

Chapter 8. The Polymerase Chain Reaction

• Introduction
• 8.1. Template and Amplification
• 8.2. Exponential Amplification
• 8.3. Polymerase Chain Reaction Efficiency
• 8.4. Calculating the Tm of the Target Sequence
• 8.5. Primers
• 8.6. Primer Tm
• 8.7. Deoxynucleoside Triphosphates
• 8.8. DNA Polymerase
• 8.9. Quantitative PCR
• Chapter Summary

Chapter 9. Real-Time PCR

• Introduction
• 9.1. The Phases of a Real-Time Polymerase Chain Reaction
• 9.2. Controls
• 9.3. Absolute Quantification by the TaqMan Assay
• 9.4. Amplification Efficiency
• 9.5. Measuring Gene Expression
• 9.6. Relative Quantification: The ΔΔCT Method
• 9.7. Relative Standard Curve Method
• 9.8. Relative Quantification by Reaction Kinetics
• 9.9. The R0 Method of Relative Quantification
• 9.10. The Pfaffl Model
• 9.11. Digital Polymerase Chain Reaction
• Chapter Summary

Chapter 10. Recombinant DNA

• Introduction
• 10.1. Restriction Endonucleases
• 10.2. Calculating the Amount of Fragment Ends
• 10.3. Ligation
• 10.4. Genomic Libraries—How Many Clones Do You Need?
• 10.5. cDNA Libraries—How Many Clones Are Enough?
• 10.6. Expression Libraries
• 10.7. Screening Recombinant Libraries by Hybridization to DNA Probes
• 10.8. Sizing DNA Fragments by Gel Electrophoresis
• 10.9. Generating Nested Deletions Using Nuclease BAL 31
• Chapter Summary

Chapter 11. Protein

• Introduction
• 11.1. Calculating a Protein's Molecular Weight From Its Sequence
• 11.2. Determining a Protein's Molecular Weight by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis
• 11.3. Protein Quantification by Measuring Absorbance at 280nm
• 11.4. Using Absorbance Coefficients and Extinction Coefficients to Estimate Protein Concentration
• 11.5. Relating Concentration in Milligrams per Milliliter to Molarity
• 11.6. Protein Quantitation Using A280 When Contaminating Nucleic Acids Are Present
• 11.7. Protein Quantification at 205nm
• 11.8. Protein Quantitation at 205nm When Contaminating Nucleic Acids Are Present
• 11.9. Labeling Proteins With Fluorescent Dyes
• 11.10. Measuring Protein Concentration by Colorimetric Assay—The Bradford Assay
• 11.11. Thin-Layer Chromatography and Rf
• 11.12. Estimating a Protein's Molecular Weight by Gel Filtration
• 11.13. Protein Volume
• 11.14. Using β-Galactosidase to Monitor Promoter Activity and Gene Expression
• 11.15. The Chloramphenicol Acetyltransferase Assay
• 11.16. Use of Luciferase in a Reporter Assay
• 11.17. DNA Polymerase Activity
• 11.18. In Vitro Translation—Determining Amino Acid Incorporation
• 11.19. The Isoelectric Point of a Protein
• Chapter Summary

Chapter 12. Centrifugation

• Introduction
• 12.1. Relative Centrifugal Force (g Force)
• 12.2. Calculating Sedimentation Times
• Chapter Summary

Chapter 13. Forensics and Paternity

• Introduction
• 13.1. Alleles and Genotypes
• 13.2. The Hardy–Weinberg Equation and Calculating Expected Genotype Frequencies
• 13.3. The Chi-Square Test—Comparing Observed to Expected Values
• 13.4. The Power of Inclusion
• 13.5. The Power of Discrimination
• 13.6. DNA Typing and a Weighted Average
• 13.7. The Multiplication Rule
• 13.8. The Paternity Index
• Chapter Summary
• More Problems

Appendix A. Using Microsoft Excel's Graphing Utility