Techniques in Bioproductivity and Photosynthesis

Introduction

Units, Symbols and Abbreviations


1. Measurement of Plant Biomass and Net Primary Production


1.1 Introduction


1.1.1 Definitions


1.1.2 Units


1.1.3 Principles


1.2 Sampling Design


1.2.1 Fully Randomized Design


1.2.2 Randomized Block Design


1.3 Measurement of Above-Ground Biomass


1.3.1 Number, Area and Shape of Quadrats


1.3.2 Harvesting


1.3.3 Sorting


1.3.4 Drying


1.3.5 Ashing


1.3.6 Determination of Energy Content


1.3.7 Carbon Analysis


1.4 Measurement of Below-Ground Biomass


1.4.1 Soil Extraction


1.4.2 Separation from Soil and Washing


1.4.3 Separation of Live Roots from Dead Matter


1.4.4 Dry Weight and Organic Weight


1.5 Non-Destructive Measurement of Biomass


1.5.1 Estimation from Stem and Leaf Dimensions


1.5.2 Remote Sensing


1.6 Estimation of Losses


1.6.1 Decomposition


1.6.2 Grazing


1.6.3 Exudation


1.7 Estimation of Net Primary Production


1.8 Experimental Work

References


2. Plant Growth Analysis


2.1 Introduction


2.2 Basic Principles


2.3 Components of Classical Growth Analysis


2.3.1 Relative Growth Rate


2.3.2 Unit Leaf Rate


2.3.3 Leaf Area Ratio


2.3.4 Specific Leaf Area and Leaf Weight Ratio


2.3.5 Leaf Area Index


2.3.6 Crop Growth Rate


2.3.7 Leaf Area Duration


2.4 Functional Growth Analysis


2.4.1 Introduction


2.4.2 The Richards Function


2.5 Experimental Investigations


2.5.1 Limitations


2.5.2 Procedure

References


3. Plant Microclimate


3.1 General Introduction


3.2 Radiation - Solar and Long Wave


3.2.1 Introduction


3.2.2 Radiation Measurements


3.3 Temperature


3.3.1 Introduction


3.3.2 Temperature Measurements


3.3.3 Use of Thermometers


3.4 Humidity


3.4.1 Introduction


3.4.2 Definitions


3.4.3 Measurements


3.5 Wind


3.5.1 Measurement


3.6 Automatic Weather Stations


3.7 Recording


3.8 Experimental Work

References


4. Canopy Structure and Light Interception


4.1 Introduction


4.2 Radiation in Canopies


4.2.1 Changes in Photon Flux Density within the Canopy


4.2.2 Considerations for Specific Canopies


4.2.3 Light Compensation


4.2.4 Measurement of Light in Canopies


4.3 Measurement of Canopy Structure


4.3.1 Leaf Area Index


4.3.2 Leaf Inclination


4.3.3 Leaf Orientation


4.3.4 Stems and Inflorescences

References


5. Water Relations


5.1 Stomatal Conductance


5.1.1 Introduction


5.1.2 Resistance or Conductance


5.1.3 Methods for Stomatal Aperture


5.1.4 Methods for Rate of Water-Vapor Loss


5.1.5 Principles and Calibration of Diffusion Porometers


5.2 Plant Water Status


5.2.1 Introduction


5.2.2 Water Content


5.2.3 Water Potential


5.2.4 Components of Water Potential


5.2.5 Pressure-Volume Curves


5.3 Soil Water Status


5.3.1 Introduction


5.3.2 Soil Water Content


5.3.3 Water Potential


5.4 Practical Work on Water Relations


5.4.1 Objectives


5.4.2 Materials


5.4.3 Procedure


5.4.4 Calculations


5.4.5 Analysis of Data


5.4.6 Suggested Additional Work

References and Further Reading


6. Measurement of CO2 Assimilation by Plants in the Field and the Laboratory


6.1 Introduction


6.1.1 The Role of CO2 Exchange Measurements


6.1.2 Measures, Symbols and Units


6.1.3 The Approach


6.1.4 Closed Systems


6.1.5 Semi-Closed Systems


6.1.6 Open Systems


6.2 Infra-Red Gas Analysis


6.2.1 Principles


6.2.2 Configurations


6.2.3 Calibration


6.3 14C Incorporation


6.3.1 Principle


6.3.2 Procedure


6.4 Measurement and Control of Gas Flow


6.4.1 Flowmetering


6.4.2 Variable-Area Flowmeters


6.4.3 Thermal Mass Flowmeters


6.4.4 Flowmeter Calibration


6.4.5 Flow Control


6.5. Chamber Conditions and Construction


6.5.1 Principles


6.5.2 Chamber Design


6.5.3 Boundary Layer Conditions


6.5.4 Temperature


6.5.5 Photon Flux Density


6.5.6 Materials


6.5.7 Air Conditioning


6.6 Analysis of Gas Exchange Measurements


6.6.1 Resistance Analogies


6.6.2 The Gaseous Diffusion Pathway


6.6.3 The A/ci Response


6.6.4 Light Response


6.7 Conclusion


6.8 Experimental Work


6.8.1 Photosynthetic Gas Exchange in the Laboratory


6.8.2 Photosynthetic Gas Exchange in the Field

References


7. Measurement of Oxygen and Chlorophyll Fluorescence


7.1 The Oxygen Electrode


7.1.1 Introduction


7.1.2 Characteristics of the Oxygen Electrode


7.1.3 Calibration


7.1.4 Other Forms of Oxygen Measurement


7.2 The Leaf Disc Electrode


7.2.1 Introduction


7.2.2 Calibration


7.2.3 Calculations Relating to Volume and Calibration


7.2.4 Experiment: Does It Work with a Leaf


7.3 Chlorophyll Fluorescence Measurement


7.3.1 Introduction


7.3.2 Principles of Measurement


7.3.3 Is Fluorescence Influenced by Carbon Assimilation


7.3.4 Fluorescence Induction


7.3.5 Complex Fluorescence Kinetics and Oscillations

References


8. Shoot Morphology and Leaf Anatomy in Relation to Photosynthesis


8.1 Shoot Morphology and the Relationship of Single Leaf to Whole Plant CO2 Assimilation and Canopy Productivity


8.2 Leaf Anatomy


8.2.1 Influence of High and Low Light Intensities on Leaf Anatomy


8.2.2 Xeromorphy


8.2.3 Leaf Anatomy Related to C3 and C4 Pathways of CO2 Fixation


8.2.4 Sub-Groupings of C4 Species


8.3 Experiments


8.3.1 Leaf Anatomy


8.3.2 Stomatal Width


8.3.3 Demonstration of 02 Evolution from Whole Plants


8.3.4 in Situ Demonstration of PSII Activity


8.3.5 Differentiation between C3 and C4 Plants by Detection of Starch In Situ


8.3.6 Demonstration of Phloem Translocation in Detached Maize Leaves

References


9. Chloroplasts and Protoplasts


9.1 Introduction


9.1.1 Plant Material


9.2 Mesophyll Protoplasts from C3, C4 and CAM Plants


9.2.1 Preparation of Leaf Tissue and the Digestion Procedure


9.2.2 Isolation and Purification


9.2.3 Storage of Protoplasts


9.2.4 Isolation of Plant Cells


9.3 Photosynthesis by Isolated Protoplasts


9.3.1 C3 Protoplasts


9.3.2 C4 Mesophyll Protoplasts


9.3.3 C4 Bundle-Sheath Strands


9.3.4 CAM Cells and Protoplasts


9.4 Chloroplast Isolation from Protoplasts


9.4.1 C3 Chloroplasts


9.4.2 C4 Chloroplasts


9.4.3 CAM Chloroplasts


9.5 Mechanical Separation of Intact Chloroplasts


9.5.1 Isolation of Chloroplasts from Spinach


9.5.2 Variations in the Procedure for Spinach


9.6 Photosynthesis by Isolated Chloroplasts


9.6.1 Criteria of Intactness


9.6.2 Intactness Assay


9.6.3 Improvement in Chloroplast Intactness


9.7 Carbon Assimilation by C3 Chloroplasts


9.7.1 Measurement of Chloroplast Photosynthesis


9.8 Materials

References


10. Photosynthetic Energy Conversion


10.1 Introduction


10.2 Measurement of Proton Flux, Photophosphorylation and Electron Transport Using Broken Chloroplasts and a pH Electrode


10.2.1 Measurement of Proton Flux


10.2.2 Measurement of Photophosphorylation


10.2.3 Measurement of Electron Transport


10.3 Partial Electron Transport Reactions Assayed with the O2 Electrode and a Conventional Recording Spectrophotometer


10.3.1 Water to Methyl Viologen


10.3.2 DCPIP to Methyl Viologen


10.3.3 Water to p-Phenylenediamine


10.3.4 Water to Silicomolybdate


10.3.5 DPC to Methyl Viologen


10.3.6 Assay for FNR Using a Recording Spectrophotometer

References


11. Carbon Metabolism


11.1 Introduction


11.2 The Carbon Reduction Cycle


11.3 Photorespiration


11.4 C4 Photosynthesis


11.5 CAM Plants


11.6 Carboxylation Enzymes


11.6.1 RUBISCO


11.6.2 Phosphoenol Pyruvate Carboxylase


11.7 Experiments


11.7.1 I4C Fixation Patterns in C, and C, Plants


11.7.2 Photorespiration


11.7.3 Assay of Carboxylases

References


12. Nitrogen Fixation


12.1 Nitrogen Metabolism


12.2 Introduction to N-Fixation


12.3 The Genus Rhizobium


12.4 Measurement of N-Fixation by Direct Means


12.5 Indirect Assay of Nitrogenase Activity


12.6 Preparation of Acetylene


12.7 Gas Chromatography


12.8 Experimental Schedule

References


13. Assimilatory Nitrate Reduction


13.1 Introduction


13.2 Enzymology


13.2.1 Nitrate Reductase


13.2.2 Nitrite Reductase


13.3 Relationship between Nitrate Reduction and Photosynthesis


13.4 Determination of Enzymatic Activities


13.4.1 Nitrate Reductase Activity


13.4.2 Nitrite Reductase Activity


13.5 Other Analytical Procedures Useful for the Study of Nitrate Assimilation


13.5.1 Determination of Nitrate


13.5.2 Chemical Determination of Ammonium and Dissolved Ammonia


13.5.3 Enzymatic Determination of Ammonium and Dissolved Ammonia


13.6 Uptake of Nitrate by Cells of Blue-Green Algae. Effects of Ammonium and MSO

References


14. Ammonia Assimilation and Amino Acid Biosynthesis


14.1 Ammonia Assimilation


14.1.1 Introduction


14.1.2 Enzyme Isolation


14.1.3 Glutamine Synthase


14.1.4 Glutamate Synthase


14.1.5 Glutamate Dehydrogenase


14.1.6 Aminotransferases


14.2 Transport of Nitrogenous Compounds


14.2.1 Compounds Utilized


14.2.2 Assay of Transport Compounds


14.2.3 Nitrogen Fixation


14.3 Biosynthesis of Amino Acids


14.3.1 The Aspartate Family


14.3.2 Selection of Amino Acid Metabolism Mutants


14.3.3 Mutagen Treatment Technique


14.3.4 Isolation and Characterization of Aspartate Kinase


14.3.5 Proline and Drought Stress

References


15. Micro-Algae: Laboratory Growth Techniques and Outdoor Biomass Production


15.1 Introduction


15.2 Growth of Micro-Algae: Techniques and Kinetics


15.2.1 Batch Culture


15.2.2 Continuous Culture


15.3 Chemostat Cultures


15.4 Synchronized Cultures


15.5 Nutrient-Limited Growth


15.6 Analytical Techniques


15.6.1 Cell Counting


15.6.2 Light Scattering (Turbidity)


15.6.3 Dry Weight Measurement


15.7 Growth, Maintenance and Preservation of Algal Cultures


15.8 Culture Media


15.9 Definition of Common Units and Terms


15.10 Technical Problems


15.11 Where to Obtain Algal Cultures


15.12 Algal Biomass Production


15.12.1 Biological Problems


15.12.2 Engineering Problems


15.12.3 Uses of Algal Biomass

References and Further Reading


16. Enzymes: Separation and Kinetics


16.1 Introduction


16.2 Enzyme Activity


16.3 Isolation of Enzymes


16.4 Protein Determination


16.5 Extraction of Proteins


16.6 Protection from Phenols and Phenol Oxidase Activity


16.7 Crude Extracts


16.8 Ammonium Sulfate Precipitation


16.9 Modern Methods of Protein Purification


16.9.1 Ion Exchange Chromatography


16.9.2 Molecular Sieve Chromatography


16.9.3 Electrophoresis


16.9.4 Isoelectric Focusing


16.10 Enzyme Kinetics


16.11 Experiments


16.11.1 Demonstration of Phenol Oxidase Activity and Design of Protective Media


16.11.2 Enzyme Purification


16.11.3 Polyacrylamide Gel Electrophoresis


16.12 The Enzyme Commission Classification System

References


17. Analytical Techniques


17.1 Protein Analysis


17.1.1 Kjeldahl Method


17.1.2 Biuret Reaction


17.1.3 Lowry Method


17.2 Analysis of Carbon Isotope Discrimination Ratios


17.3 Measurement of Light Absorption


17.3.1 The Absorption of Light by Molecules


17.3.2 Experimental Method


17.4 Chlorophyll Determination


17.4.1 Chlorophyll in Higher Plants


17.4.2 Algal Chlorophyll


17.5 Measurement of Starch and Sucrose in Leaves


17.5.1 Preparation of Leaf Extracts for Metabolite Measurements


17.5.2 Measurement of Sucrose


17.5.3 Measurement of Starch


17.6 Mineral Analysis of Plants, Soil and Water

References

Appendices

A. Equipment for Field and Laboratory Studies of Whole Plant and Crop Photosynthesis and Productivity Research

B. Experimental Design and Presentation of Results

C. Biomass Production and Data

D. Conversion Factors and Approximate Conversion Factors

E. Solar Radiation on the Earth

Index