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OPPORTUNITI
B10
IN
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Committee on Research Opportunities in Biology
Board on Biology
Commission on Life Sciences
National Research Council
NATIONAL ACADEMY PRESS
Washington, D.C. 1989
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COMMITTEE ON RESEARCH OPPORTUNITIES
IN BIOLOGY
PETER H. RAVEN (Chairman), Missouri Botanical Garden, St. Louis
CHARLES R. CANTOR, Columbia University College of Physicians and
Surgeons, New York, New York
W. MAXWELL COWAN, Howard Hughes Medical Institute, Bethesda, Maryland
JAMES D. EBERT, Johns Hopkins University, Baltimore, Maryland
THOMAS EISNER, Cornell University, Ithaca, New York
GERALD R. FINK, Whitehead Institute and Massachusetts Institute of
Technology, Cambridge
WILFORD R. GARDNER, University of California, Berkeley
DANIEL HARTL, Washington University School of Medicine, St. Louis,
Missouri
LEROY HOOD, California Institute of Technology, Pasadena
ERNEST G. JAWORSKI, Monsanto Company, St. Louis, Missouri
ERIC R. KANDEL, Columbia University College of Physicians and Surgeons and
Howard Hughes Medical Institute, New York, New York
HANS KENDE, Michigan State University, East Lansing
GEORGE R. PALADE, Yale University School of Medicine,
New Haven, Connecticut
MARY-LOU PARDUE, Massachusetts Institute of Technology, Cambridge
WILLIAM E. PAUL, National Institutes of Health, Bethesda, Maryland
THOMAS D. POLLARD, Johns Hopkins Medical School, Baltimore, Maryland
RUSSELL ROSS, University of Washington School of Medicine, Seattle
MELVIN I. SIMON, California Institute of Technology, Pasadena
DONALD F. STEINER, University of Chicago, Illinois
EDWARD O. WILSON, Harvard University, Cambridge, Massachusetts
NRC Staff
JOHN E. BURRIS, Study Director
CLIFFORD J. GABRIEL, Senior Program Officer
DAVID J. POLICANSKY, Senior Program Officer (through December 1986)
WALTER G. ROSEN, Senior Program Officer (through December 1986)
CAITILIN GORDON, Editor
M. FRANCES WALTON, Administrative Secretary (through August 1988)
KATHY L. MARSHALL, Senior Secretary
· . ~
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Panel on Cell Organization
THOMAS POLLARD (Co-chairman), Johns Hopkins Medical School, Baltimore,
Maryland
BRUCE ALBERTS (Co-cha~rman), University of California, San Francisco
THOMAS DEUEL, Jewish Hospital at Washington University Medical Center,
St. Louis, Missouri
LELAND HARTWELL, University of Washington, Seattle
PHILLIP SHARP, Massachusetts Institute of Technology, Cambridge
Panel on Ecology and Ecosystems
SIMON LEVIN (Chairman), Cornell University, Ithaca, New York
JAMES EHLERINGER, University of Utah, Salt Lake City
THOMAS EISNER, Cornell University, Ithaca, New York
STEPHEN HUBBELL, University of Iowa, Iowa City
HOLGER JANNASCH, Woods Hole Oceanographic Institution, Woods Hole,
Massachusetts
PETER RAVEN, Missouri Botanical Garden, St. Louis
THOMAS SCHOENER, University of California, Davis
PETER V1TOUSEK, Stanford University, Stanford, California
Panel on Evolution and Diversity
DANIEL HARTL (Co-chairr~n), Washington University School of Medicine,
St. Louis, Missouri
MICHAEL CLEGG (Co-chairman), University of California, Riverside
J. WILLIAM SCHOPF, University of California, Los Angeles
DOUGLAS FUTUYMA, State University of New York at Stony Brook
DAVID RAUP, University of Chicago, Illinois
EDWARD WILSON, Harvard University, Cambridge, Massachusetts
CARL WOESE, University of Illinois, Urbana
Panel on Genome Organization and Expression
GERALD FINK (Co-chairman), Whitehead Institute and Massachusetts Institute of
Technology, Cambridge
JOSEPH GALL (Co-chcurman), Carnegie Institution of Washington, Baltimore,
Maryland
PETER QUAIL, Plant Gene Expression Center, Albany, California
MELVIN SIMON, California Institute of Technology, Pasadena
JOAN STERN, Yale University School of Medicine, New Haven, Connecticut
RAYMOND WHITE, Howard Hughes Medical Institute and University of Utah
Medical School, Salt Lake City
IV
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Panel on Growth and Development
MARY-LOU PARDUE (Co-cha~rman), Massachusetts Institute of Technology,
Cambridge
MARC KIRSCHNER (Co-chairman), University of California Medical School,
San Francisco
SUSAN BRYANT, University of California, Vine
COREY GOODMAN, University of California, Berkeley
PHILIPPA MARRACK, National Jewish Hospital, Denver, Colorado
DAVOR SOLTER, Wistar Institute, Philadelphia, Pennsylvania
Panel on Immune System, Pathogens, and Host Defenses
WILLIAM PAUL (Co-chairman), National Institutes of Health, Bethesda,
Maryland
BERNARD FIELDS (Co-chairman), Harvard Medical School, Boston,
Massachusetts
ZANVIL COHN, Rockefeller University, New York, New York
STANLEY FALKOW, Stanford University, Stanford, California
MALCOLM GEFTER, Massachusetts Institute of Technology, Cambridge
DAVID SACHS, National Institutes of Health, Bethesda, Maryland
MATTHEW SCHARFF, Albert Einstein College of Medicine, New York,
New York
THOMAS WALDMANN, National Institutes of Health, Bethesda, Maryland
Panel on Integrative Approaches to Organism Function and Disease
RUSSELL ROSS (Chairman), University of Washington, Seattle
HENRY BOURNE, University of California, San Francisco
MICHAEL CZECH, University of Massachusetts Medical School, Worcester
FRED FOX, University of California, Los Angeles
BERTIL HILLE, University of Washington School of Medicine, Seattle
PHILIP NEEDLEMAN, Washington University, St. Louis, Missouri
ALEXANDER NICHOLS, University of California, Berkeley
Panel on Molecular Structure and Function
CHARLES CANTOR (Co-chairman), Columbia University College of Physicians
and Surgeons, New York, New York
FREDERICK RICHARDS (Co-chairman), Guilford, Connecticut
MARK PTASHNE, Harvard University, Cambridge, Massachusetts
LUBERT STAYER, Stanford Medical School, Stanford, California
NIGEL UNWIN, Stanford University, Stanford, California
DONALD WILEY, Harvard University, Cambridge, Massachusetts
v
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Panel on Neurobiology and Behavior
W. MAXWELL COWAN (Co-chairman), Howard Hughes Medical Institute,
Bethesda, Maryland
ERIC KANDEL (Co-chairman), Columbia University College of Physicians and
Surgeons and Howard Hughes Medical Institute, New York, New York
ALBERT AGUAYO, Montreal General Hospital, Canada
EMILIO BlZ/.l, Massachusetts Institute of Technology, Cambridge
JOHN DOWLING, Harvard University, Cambridge, Massachusetts
GERALD FISCHBACH, Washington University School of Medicine, St. Louis,
Missouri
BERT HOLLDOBLER, Harvard University, Cambridge, Massachusetts
MICHAEL POSNER, University of Oregon, Eugene
SOLOMON SNYDER, Johns Hopkins University School of Medicine, Baltimore,
Maryland
Panel on New Technology and Industrial Biotechnology
ERNEST JAWORSKI (Co-chairman), Monsanto Company, St. Louis, Missouri
LEROY HOOD (Co-chairman), California Institute of Technology, Pasadena
RITA COLWELL, University of MaIyland, College Park
CHARLES COONEY, Massachusetts Institute of Technology, Cambridge
ROBERT KAMEN, Genetics Institute, Cambridge, Massachusetts
ROBERT SHULMAN, Yale University, New Haven, Connecticut
Panel on Plant Biology and Agriculture
HANS KENDE (Co-chairman), Michigan State University, East Lansing
WILFORD GARDNER (Co-chairman), University of California, Berkeley
JOHN BOYER, University of Delaware, Lewes
TSUNE KOSUGE (deceased), University of California, Davis
SHARON LONG, Stanford University, Sanford, California
DONALD ORT, USDA-AAS-University of Illinois, Urbana
Vl
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BOARD ON BIOLOGY
FRANCISCO J. AYALA (Chairman), University of California, Irvine
NINA V. FEDOROFF, Carnegie Institution of Washington, Baltimore, Maryland
TIMOTHY H. GOLDSMITH, Yale University, New Haven, Connecticut
RALPH W. F. HARDY, Boyce Thompson Institute of Plant Research, Ithaca,
New York
ERNEST G. JAWORSKI, Monsanto Company, St. Louis, Missouri
SIMON A. LEVIN, Cornell University, Ithaca, New York
HAROLD A. MOONEY, Stanford University, Stanford, California
HAROLD J. MOROW1'I Z. George Mason University, Fairfax, Virginia
MARY-LOU PARDUE, Massachusetts Institute of Technology, Cambridge
WILLIAM E. PAUL, National Institutes of Health, Bethesda, Maryland
DAVID D. SABAT1NI, New York University, New York
MICHAEL E. SOULE, University of Michigan, Ann Arbor
MALCOLM S. STEINBERG, Princeton University, Princeton, New Jersey
DAVID B. WAKE, University of California, Berkeley
BRUCE M. ALBERTS (0~-officio), University of California, San Francisco
NRC Staff
JOHN E. BURRIS, Executive Director, Commission on Life Sciences
· .
v``
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COMMISSION ON LIFE SCIENCES
BRUCE M. ALBERTS (Chairman), University of California, San Francisco
PERRY L. ADKISSON, The Texas A&M University System, College Station,
Texas
FRANCISCO J. AYALA, University of Califomia, Irvine
J. MICHAEL BISHOP, University of California Medical Center, San Francisco
FREEMAN J. DYSON, Institute for Advanced Study, Princeton, New Jersey
NINA V. FEDOROFF, Carnegie Institution of Washington, Baltimore, Maryland
RALPH W. F. HARDY, Boyce Thompson Institute of Plant Research, Ithaca,
New York
RICHARD J. HAVEL, University of California School of Medicine,
San Francisco
LEROY E. HOOD, California Institute of Technology, Pasadena
DONALD F. HORNIG, Harvard School of Public Health, Boston, Massachusetts
ERNEST G. JAWORSKI, Monsanto Company, St Louis, Missouri
SIMON A. LEVIN, Cornell University, Ithaca, New York
HAROLD A. MOONEY, Stanford University, Stanford, California
STEVEN P. PAKES, University of Texas Southwestern Medical School, Dallas
JOSEPH E. RALL, National Institutes of Health, Bethesda, Maryland
RICHARD D. REMINGTON, University of Iowa, Iowa City
PAUL G. RISSER, University of New Mexico, Albuquerque
RICHARD B. SETLOW, Brookhaven National Laboratory, Upton, New York
TORSTEN N. WIESEL, Rockefeller University, New York, New York
NRC Stay
JOHN E. BURRIS, Executive Director
.. .
Vli!
OCR for page R9
Acknowledgments
The committee would like to acknowledge the thoughtful work of the contribu-
tors and reviewers:
Edward Adelberg
Bruce Baker
Edwin Beachey
Roger Beachy
Hans Bode
Lawrence Bogorad
Marianne Bonner-Fraser
Dan Brower
Bob Buchanan
Guy Bush
Judith Campbell
Thomas Cech
Robert Chanock
Bruce Chesebro
Tom Cline
C. Robert Cloninger
R. John Collier
Robert J. Collier
John Collins
Jonathon Cooke
George Cross
Kathryn Crossin
Deborah Delmer
William Earnshaw
Robert Fraley
Michael Freeling
Larry Gerace
Norton Gilula
Timothy Goldsmith
Leslie Gottlieb
Antonio Gotto
J. Frederick Grassle
Paul Green
Douglas Hanahan
Richard Harlan
George Haughn
Ari Helenius
Samuel Hellman
John Hildebrand
H. Robert Horvitz
Rudolf Jaenisch
~y Kahn
Arthur Kelman
Eric Knudsen
Thomas Kornberg
Stephen Krane
~x
Michael Levine
Jane Lubchenco
Harry MacWiD~s
Thomas Maniatis
Lynn Margulis
Paul Marks
David Martin
Gail Martin
Victor McKusick
Douglas Melton
Harold Mooney
J. Anthony Movshon
Howard Nash
June Nasrallah
Daniel Nathans
Stephen O'Brien
William Ogren
Michael Oldstone
Gordon Orians
David Page
David Pisetsky
Peter Quail
Calvin Quate
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ACKNOWLEl)GME7VTS
Richard Root
Michael Rosenzweig
John Roth
Erkki Ruoslahti
Frank Ruddle
Jay Savage
Luis Sequeira
David Schlessinger
James Schwartz
Matthew Scott
Michael Sheetz
Emil Skamene
Allan Spradling
Malcolm Steinberg
William Sugden
James Tavares
D. Lansing Taylor
Howard Temin
Samuel Thier
Robert Trelstad
David Van Essen
Graham Walker
David Wake
Allan Wilson
Thomas Woolsey
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Preface
In 1970 the National Research Council published Biology and the Future of
Man. This report, edited by Philip Handler, then president of the National Acad-
emy of Sciences, summarized the state of biology at that time. Now, almost 20
years later, the National Research Council, with the publication of this report,
reevaluates research opportunities in biology and attempts to convey the current
excitement in the field of biology. Our report differs from Biology and the Future
of Man because of the enormous advances that have occurred in biology over the
past two decades. The field has, in fact, changed to the point that no single
individual can hope to grasp all of the new activities and opportunities.
To address this daunting task, a committee of distinguished scientists began
in late 1985 to determine the major research areas that exist in the field of biology
and then to discuss how advances in each of these areas can be maximized and
how and where possible interactions among biologists of various subdisciplines
and biologists and other scientists can be facilitated to lead to new interdiscipli-
nary insights and approaches. This committee of 20 individuals soon realized that
such a goal would require the assistance of many other experts. Toward this end
the committee organized 11 panels, each with at least one individual from the
committee. Each panel was asked to produce a report of about 50 pages stressing
the current and future opportunities for exciting research in their area of expertise
and also to stress the interconnections of biological scientists with others from
different areas of biology and from other disciplines. Even the panels found that
they needed assistance; thus the input of additional individuals was solicited for
the report.
The steering committee edited these reports, in some cases combining the
efforts of several groups to produce a single chapter, in other instances letting
panel topics stand as chapters in the final report. The committee report then
OCR for page R12
. .
xzz
PREFACE
underwent an extensive review process. The reviewers' comments were ex-
tremely useful to the committee in polishing and producing the final draft of the
report.
This report reflects the panoply of interesting and exciting topics that consti-
tute biology. Some areas are barely covered and still others not at all. This is by
necessity: There is just too much information today in biology, a fact illustrated
by the length of high school and college introductory textbooks that can character-
istically exceed 1,000 pages. In selecting areas to be included we chose those that
represent major themes in biological research. In many cases such selections
were difficult, since most of biology today is expanding as new techniques and
ideas are applied to old fields or to new areas.
We have produced this volume for a large audience: biologists; policymakers
both in government, universities, and in industry; and other scientists from a
variety of disciplines who may interact with biologists. We hope that each of
these groups will learn from this book. For all of you who read this report, our
goal is to leave you with some understanding and appreciation of the diversity of
problems and opportunities that await the biologist. Some of these opportunities
will provide us with a better understanding of the basic workings of life, and
others will have immediate application in our lives through medicine, agriculture,
or environmental management.
In any such effort it is important to acknowledge the work of the many
individuals, in addition to the committee members, who contributed to this effort.
Those who served on the panels and contributed material to the panels are listed in
the section following the committee list. My thanks to all of them. I also thank
the staff of the National Research Council for their efforts. Frances Walton
cheerfully provided the administrative support necessary for the committee to
meet and function. Kathy Marshall spent long hours skillfully preparing the many
drafts of the manuscript. Caitilin Gordon provided expert editorial assistance.
Walt Rosen and David Policansky provided panel support early in the project and
continued to contribute as the project progressed. In particular, I acknowledge the
dedicated staff support of John Burris and Cliff Gabriel. John was the project
director throughout the effort, organizing the committee and helping it through
the early drafts. Cliff shepherded the effort through its later stages, assuming
oversight of the manuscript in later drafts and the review process, a labor of great
effort and dedication. To them, and all others who worked so hard to produce this
study, I offer my most sincere appreciation.
PETER H. RAVEN, Chairman
Committee on Research
Opportunities in Biology
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Contents
EXECUTIVE SUMMARY
1 THE NEW BIOLOGY
2 NEW TECHNOLOGIES AND INSTRUMENTATION
RECOMBINANT DNA TECHNIQUES, 19
Transformation of Higher Organisms, 21; Making a Transgenic
Animal, 23; Creating Transgenic Plants, 26; Future Prospects, 26
MONOCLONAL ANIIBODES, 27
MICROCHEMICAL TECHNIQUES, 28
FLOW CYTOMETRY, 31
MICROSCOPY, 32
Video-Enhanced Contrast Microscopy, 33; Low-Light-Dose
Microscopy, 33; Scanning Acoustic Microscope, 34; Scanning
Tunneling and Atomic Force Microscope, 34
MAGNETIC RESONANCE, 35
COMPUTERS AND DATA ANALYSIS, 36
BIOLOGY AND THE FUTURE, 37
15
19
3 MOLECULAR STRUCTURE AND FUNCTION 39
PRIMARY STRUCTURE, 43
THREE-DIMENSIONAL STRUCTURE, 51
MOLECULAR ASSEMBLES, 60
DIRECTED MODIFICATION OF PROTEINS, 67
FOLDING, 71
NEW TECHNIQUES AND INSTRUMENTATION, 75
. . ~
Ill
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All
4 GENES AND CELLS ........................
RESEARCH STRATEGIES, 78
THE NUCLEUS, 79
Nuclear Envelope, 80; Chromosomes, 81; The Nucleolus, 84
GENES AND GENE ACTION, 84
Genetic Analysis, 85; The Genome, 88; DNA Replication, 90;
Recombination, 93; DNA Repair and Mutagenesis, 96; Gene
Expression, 98; Genome Organization, 103
CYTOPLASM: ORGANELLES AND FUNCTIONS, 104
Protein Synthesis and Regulation, 105
MITOCHONDRIA: FUNCTION AND BIOGENESIS, 108
CELL MOTILITY AND THE CYTOSKELETON, 111
Cell Structure, 111; Cell Movement, 113
CELL MEMBRANE, 115
THE EXTRACELLULAR MATRIX OF ANIMALS, 117
CELL REGULATION, 118
Cell Division, 118; Cell-to-Cell Communication, 120; Growth
Factors, 122; Receptors, 123; Transmembrane Signaling, 124;
Oncogenes, 133
GENERAL PLANT CELL BIOLOGY, 134
Chloroplasts, 135; The Plant Cell Wall, 137
5 DEVEL()PMENT
COlVTENTS
77
.......... 140
DEVELOPMENT BEGINS WITH GAMETOGENESIS, 142
CELL DIVISION, GROWTH, AND DEVELOPMENTAL
TIMING, 146
DlFFiiRENTL\L GENE EXPRESSION, 153
CELL MOVEMENT AND CELL ADHESION, 159
POSITIONAL INFORMATION, 164
DEVELOPMENT IS FOR ADULT ANIMALS TOO, 167
SPECIAL PROBLEMS IN PLANT DEVELOPMENT, 169
Plant Cell Growth, 171; The Development of Plant Organs, 174
6 THE NERVOUS SYSTEM AND BEHAVIOR ....
NERVE CELL COMMUNICATION, 182
THE CYTOSKELETON AND TO TRANSPORT OF
MATERIALS WITH NERVE CELLS, 187
Slow Axonal Transport, 188; Rapid Axonal Transport, 189
DEVELOPMENT OF THE NERVOUS SYSTEM, 189
Selective Cell Aggregation, 192
NEURAL PLASTICITY AND ELEMENTARY FORMS OF
LEARNING, 198
175
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CONTENTS
NEUROBIOLOGY OF PERCEPTION: VISION, 201
Behavioral Studies of Vision in Animals, 204
NEUROBIOLOGY OF MOTOR CONTROL, 205
NEUROBIOLOGY OF COGNITION, 210
Attention, 212; Psychobiology of Development, 213
BEHAVIORAL ECOLOGY, 214
ABNORMALITIES OF BEHAVIOR, 219
7 THE IMMUNE SYSTEM AND INFECTIOUS
DISEASES . e e e e e e e e e e e · e e e e e e e e e · e e e · e · e e e e e e e e e e e e · e
THE IMMUNE SYSTEM, 224
Specificity of the Immune Response, 225; B Lymphocytes, 229;
T Cells, 233; Regulation of the Immune Response, 239;
Complement and Other Effector Molecules, 241; Hypersensitivity,
Inflammation, and Phagocytosis, 241
INFECTIOUS DISEASES, 244
Entry into the Host, 245; Spread in the Host, 249; Interactions of
Kineses with the Host Cell, 251; Cell and Tissue Tropism, 252;
How Is Injury Mediated?, 254; Persistent Infections, 258
CONCLUSION, 258
xv
-224
8 EVOLUTION AND DIVERSITY 260
THE EVOLUTIONARY PROCESS, 265
THE RESULT OF EVOLUTION, 270
EVOLUTIONARY HISTORY, 279
CURRENT STATUS OF RESEARCH, 283
Collections and Special Facilities, 284
9 ECOLOGY AND ECOSYSTEMS 287
INTRODUCTION, 287
IDEAS AND APPROACHES IN ECOLOGY, 288
The Responses of Organisms to Environmental Variations, 288;
Structure and Regulation of Populations, 290; Chemical
Ecology, 292; Behavioral Ecology, 295; Genetic Ecology, 298;
The Union of Behavioral, Population, and Genetic Ecology, 299;
Ecosystem and Community Ecology, 301
HUMAN-CAUSED ENVIRONMENTAL CHANGES, THE
PROBLEMS THEY CAUSE, AND SOME SOLUTIONS, 305
Species Loss and Conservation Ecology, 307; Global Climate
Change, 313
TECHNOLOGICAL AND METHODOLOGICAL
ADVANCES, 314
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XVI
i
Remote Sensing, 314; Analytic Chemistry, 315; Tools for
Studying Paleoecology, 315; Stable Isotopes, 316;
Biotechnology, 317; Models in Ecology, 317
CONCLUSION, 321
10 ADVANCES IN MEDICINE,- THE BIOCHEMICAL
PROCESS INDUSTRY, AND ANIMAL
AGRICULTURE . e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e e
ADVANCES IN MEDICINE, 323
Molecular Pharmacology and Human Disease, 324; New
Approaches to Understanding Health and Disease: The
Lipoproteins, 328
ARTHRITIS, 334
CANCER: CURRENT STATUS AND SOME NEW
APPROACHES TO ITS CONTROL, 336
Cancer Therapy, 337; Newer Approaches to Cancer Control, 339
ADVANCES IN UNDERSTANDING HUMAN GENETIC
DISORDERS, 344
Elucidation of the Primary Genetic Defect, 345; Gene
Diagnosis, 346; Gene Therapy, 347
NEW APPROACHES FOR CONTROL OF MICROBIAL
INFECTION, 348
Vaccines, 349; Antiviral Agents, 351
ADVANCES IN THE BIOCHEMICAL PROCESS INDUSTRY, 352
Goals in Industrial Biotechnology, 353
ADVANCES IN ANIMAL AGRICULTURE, 359
Growth Regulation, 360; Reproduction, 361; Lactation, 362;
Infectious Disease, 362; Production of Feedstocks, 363; Livestock
Improvement, 363
11 PLANT BIOLOGY AND AGRICULTURE...............
PLANTS AND THEIR ENVIRONMENT, 366
PHOTOSYNTHESIS, 371
NITROGEN FIXATION, 377
PLANT GROWTH AND DEVELOPMENT, 381
Plant Hormones, 381; Environment, 384; Plant Reproduction, 385
PLANT-PATHOGEN INTERACTIONS, 390
GENETIC IMPROVEMENT OF PLANTS, 397
Tissue Culture, 397; Plant Cell Transformation, 400
CONTENTS
323
365
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CONTENTS
12 BIOLOGY RESEARCH INFRASTRUCTURE AND
RECOMMENDATIONS ..............................
TRAINING, 405
Women and Minondes, 409; Current Training Needs, 409
EMPLOYMENT, 412
LABORATORY COSTS AND EQUIPMENT NEEDS, 414
FUNDING, 416
LARGE DATA BASES AND REPOSITORIES, 420
RESEARCH CENTERS AND THE INDIVIDUAL
INVESTIGATOR, 421
REFERENCES AND NOTES, 422
INDEX
XVt!
403
425
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OPPORTUNITIES IN
BIOLOGY
-
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