Lab Manual 2022 1

Chapter 1: Microscopy and the Cell

Objectives

  • Study cell structure and characteristics of different types of cells.

    • Identify organelles: nuclei, chloroplasts, and mitochondria.

    • Learn which organelles are not visible using a light microscope.

  • Become familiar with the light microscope.

Cell Structure

  • Understanding cell structure is fundamental to biology.

  • Cell Theory:

    1. All organisms are made up of cells.

    2. All cells arise from other cells.

Prokaryotic and Eukaryotic Cells

  • Differences in cell organization:

    • Prokaryotic Cells:

      • Lack a true nucleus and membrane-bound organelles.

      • DNA is located in the nucleoid, forming a single, circular chromosome.

      • Do not divide by mitosis.

    • Eukaryotic Cells:

      • Contain a true nucleus with a nuclear envelope.

      • Have multiple linear chromosomes with histones and proteins.

      • Divide through mitosis and contain membrane-bound organelles (e.g., mitochondria, chloroplasts).

Endosymbiosis Theory

  • Mitochondria and chloroplasts contain DNA similar to prokaryotic DNA.

  • Theory states these organelles originated from free-living bacteria that entered a symbiotic relationship with early eukaryotic cells.

Table 1-1: Differences Between Prokaryotic and Eukaryotic Cells

Characteristic

Prokaryotic Cells

Eukaryotic Cells

Membrane-bound organelles

None

Mitochondria, Chloroplasts, Golgi, etc.

Nucleus

No true nucleus (nucleoid present)

True nucleus

Organization of DNA

Single circular chromosome

Multiple linear chromosomes

Type of cell division

No mitosis

Mitosis

Size

Small

Large

Types of Prokaryotic Organisms

  • Includes:

    • Archaebacteria (methanogenic, reduce sulfur)

    • Pseudobacteria (tiny, no true cell walls)

    • Bacteria (true cell walls)

    • Cyanobacteria (blue-green algae)

Characteristics of Prokaryotic Cells

  • Autotrophic and heterotrophic modes of nutrition.

  • Play important roles in nutrient cycling (saprophytic bacteria degrade organic materials).

Prokaryotic Cell Structure

  • Consist of cytoplasm, cell membrane, and cell wall.

  • DNA located in the nucleoid; may have a capsule for protection and flagella for locomotion.

Eukaryotic Organisms

  • Include protists, fungi, plants, and animals.

Protists

  • Simplest eukaryotic organisms.

  • Mostly unicellular; some colonial; diverse in nature.

Protozoa

  • Diverse group; mostly unicellular and motile; lack cell walls.

  • Types examined:

    • Amoeba: Moves via pseudopodia; some are parasitic.

    • Paramecium: Ciliated organisms with specialized nuclei for reproduction.

Algae

  • Photosynthetic; range from single cells to large colonies.

  • Example: Euglena: photosynthetic but lacks a true cell wall.

Locomotion in Protists

  • Amoeba: pseudopodia.

  • Paramecium: cilia.

  • Euglena: flagella.

Multi-Cellular Organisms

  • Include fungi, plants, and animals.

  • Evolution of multicellularity allows for specialization in tissues and organs.

Plant Cells

  • Autotrophic, have cell walls, chloroplasts, and central vacuoles.

Observations

  • Observations of various plant cells like Elodea, onion, and potato cells for structures (e.g., chloroplasts, vacuoles).

Animal Cells

  • Lack cell walls and chloroplasts, present in various forms such as cheek cells.

Chapter 2: Properties of Biological Membranes: Osmosis/Diffusion

Membranes

  • Surround all cells; define boundaries and regulate internal environment.

  • Composed of lipids, proteins, and carbohydrates.

Functions of Cell Membranes

  1. Barrier: Blocks entry of most substances.

  2. Transport: Allows movement of metabolites; includes passive transport (diffusion and osmosis) and active transport.

  3. Organization: Helps organize cellular processes.

  4. Communication: Coordinates activities through receptor binding.

  5. Recognition: Allows cells to identify each other for immune response.

Diffusion and Osmosis

  • Diffusion: Movement from high to low concentration; vital for nutrient and waste exchange.

    • Osmosis: Specific to water movement through semipermeable membranes.

  • Types of solutions:

    • Hypotonic (cell swells)

    • Isotonic (no change)

    • Hypertonic (cell shrinks)

Lab Procedures

  • Experiments designed to demonstrate concepts of osmosis using dialysis tubing.

  • Measuring sizes of cells in various solutions and observing implications of osmotic pressure on cell integrity.

Chapter 3: DNA Isolation from Plant Nuclei and Human Cheek Cells

Summary

  • Focus on isolating DNA from eukaryotic cells via disruption and centrifugation.

Procedure Overview

  • Use of detergents (SDS) and protease to break open cells and inhibit DNA-degrading enzymes.

  • Steps to extract DNA, including precipitation with ethanol.

Chapter 4: Introduction to DNA Fingerprinting

Structure of DNA

  • Overview of nucleotides and double helix structure.

Restriction Enzymes

  • Role in cutting DNA at specific sequences.

  • EcoRI example and how it generates DNA fragments.

Laboratory Techniques

  • Proper pipetting and digestion of DNA samples.

  • Usage of color-coded tubes for sample identification.

Chapter 5: DNA Fingerprinting II-Electrophoresis and Analysis of DNA Samples

Gel Electrophoresis

  • Technique for separating DNA fragments based on size.

Steps

  1. Prepare agarose gel and load samples.

  2. Apply electric current to separate fragments.

  3. Analyze migration patterns to draw conclusions about DNA sources.