Plant Tissue Culture. Techniques and Experiments

172 Pages·1992·4.257 MB·English

by  Robert H. Smith (Eds.)

Plait Tissue Culture
T e c h n i q u es a nd E x p e r i m e n ts
by
ROBERTA H. SMITH
Department of Soil and Crop Sciences
Texas Agricultural Experiment Station
Texas A & M University
College Station, Texas
ACADEMIC PRESS, INC.
Harcourt Brace Jovanovich, Publishers
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Copyright © 1992 by ACADEMIC PRESS, INC.
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United Kingdom Edition published by
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Plant Tissue Culture: Techniques and Experiments
by Roberta H. Smith
International Standard Book Number 0-12-650340-0
PRINTED IN THE UNITED STATES OF AMERICA
92 93 94 95 96 97 KP 9 8 7 6 5 4 3 2 1
To my parents
Preface
This manual resulted from the need for plant tissue culture
laboratory exercises that demonstrate major concepts and
that use plant material that is available year round. Each
chapter on the main plant cell culture techniques has plant
material options for selecting the explant source, including
seed catalogs, plant nurseries, feed stores, landscape plant
materials, and produce departments at grocery stores. In
short, the strategy in developing this manual has been to de-
vise exercises that do not require maintenance of an exten-
sive collection of plant materials, and to give students the
opportunity to work with such materials. In that respect, this
manual is different from those focusing on carrot or tobacco
cell culture systems.
The exercises in this manual have been used by a wide
range of students, from high school (science fair and 4-H
projects) to undergraduate and graduate levels. The manual
is predominantly directed at upper-level college or university
classes in which the students have taken courses in chemis-
try, plant anatomy, and plant physiology.
Before starting the exercises the student should exam-
ine Chapters 1-3, which deal with the setup of a tissue cul-
ture laboratory, media preparation, and explants and aseptic
XI
xii Preface
technique. The information in these chapters will be needed
in the exercises that follow.
The brief introduction to each chapter is not intended to
be a review of the chapter's topic but rather a complement
to lecture discussions of the topic.
In many instances, plant material initiated in one exercise
is used in subsequent exercises. Refer to Scheduling and
Interrelationships of Exercises to obtain information on the
time required to complete the exercises and how they relate
to one another.
All of the exercises have been successfully accomplished
for at least two semesters, and most have been successfully
used for five semesters. Tissue culture, however, is still more
art than science, and variation in individual exercises can be
expected.
Roberta H. Smith
Attainments
I would like to acknowledge the teaching assistants who
helped in developing some of these exercises: Daniel
Caulkins, Cheryl Knox, John Finer, Richard Norris, Ann Reilley-
Panella, Ricardo Diquez, Eugenio Ulian, Shelley Gore, Sara
Perez-Ramos, Jeffrey Callin, and Greg Peterson. Additionally,
all the students who have taken my tissue culture course
since 1979 have been important in developing and improv­
ing these exercises. Kay Hesby is acknowledged for proofread­
ing the manuscript. Linda Kocman, in particular, was
important in the proofreading, formatting, and typing of the
many versions of this manual.
XIII
'ίιιΐιπΐιιΐιιι;: and
Interrelationships of Exercises
I. Aseptic Germination of Seed (Chapter 3)
Carrot: 1-2 wk; Cotton, Sunflower: 1 wk
A. Callus Induction (Chapter 4): 6 wk
Broccoli, Lemon: 6 wk
Carrot: 2 subcultures, 6 wk each = 3 mo
1. Salt Selection in Vitro (Chapter 4): 4 wk
2. Suspension Culture (Chapter 5): 2 wk
Carrot
a. Somatic Embryogenesis (Chapter 5): 3-4 wk
b. Explant Orientation (Chapter 4): 6 wk
Cotton: 2 subcultures, 6 wk each
1. Protoplast (Chapter 10): 2 days
2. Cellular Variation (Chapter 4): 4 wk
3. Growth Curves (Chapter 4): 6 wk
II. Tobacco Seed Germination (Chapter 4): 3 wk
A. Callus Induction (Chapter 4): 2 subcultures, 6 wk
each
xv
xvi Scheduling and Interrelationships of Exercises
III. Establishment of Competent Cereal Cell Cultures
(Chapter 4): 2-3 wk
A. Rice Subculture (Chapter 5): 3 wk
1. Plant Regeneration: 4 wk
IV. Potato Shoot Initiation (Chapter 5): 6 wk
A. Potato Tuberization (Chapter 5): 4-6 wk
V. Douglas Fir Seed Germination (Chapter 3): 2-4 wk
A. Primary Morphogenesis (Chapter 5): 4 wk
VI. Petunia Leaf Disk Transformation (Chapter 11): 6 wk
VII. Petunia Shoot Apex Transformation (Chapter 11): 4-6 wk
VIII. Solitary Exercises
A. Bulb Scale Dormancy (Chapter 5): 6-8 wk
B. Datura Anther Culture (Chapter 6): 4-8 wk; 10 wk
to obtain flowering plants
C. African Violet Anther Culture (Chapter 6): 7-8 wk
D. Tobacco Anther Culture (Chapter 6): 7-8 wk;
2-3 mo to obtain flowering plants
E. Corn Embryo Culture (Chapter 7): 72 hr
F. Crabapple and Pear Embryo Culture (Chapter 7): 2-3 wk
G. Shoot Apical Meristem (Chapter 8): 4-6 wk
H. Diffenbachia Meristem (Chapter 8): 4-6 wk
I. Garlic Propagation (Chapter 8): 4 wk
J. Boston Fern Propagation (Chapter 9)
Stage I: 6-8 wk
Stage II: 4-6 wk
Stage III: 2-3 wk
K. Staghorn Fern Propagation (Chapter 9)
Stage I: 2-3 wk
Stage II: 6 wk
Stage III: 4-6 wk
Scheduling and Interrelationships of Exercises xvil
L. Ficus Propagation (Chapter 9)
Stage I: 4-6 wk
Stage II: 4-6 wk
Stage III: 4 wk
M. Kalanchoe Propagation, Stages I & II (Chapter 9): 4 wk
N. Rose Propagation, Stages I & II (Chapter 9): 8 wk
0. African Violet, Stages I & II (Chapter 9): 4 wk
P. Pitcher Plant, Stages I & II (Chapter 9): 6 wk
Q. Cactus Propagation (Chapter 9)
Stage I: 4-6 wk
Stage II: 4-6 wk
Stage III: 8 wk
D
Setup of a Tissue Culture
Laboratory
The tissue culture area should be kept picked up and clean
at all times. This is importan t to ensure clean cultures and
reproducible results. Avoid having potted plants in this area
becaus e they can be a source of mites and other contami -
nating organisms . Avoid field or greenhous e work immediatel y
before entering the laborator y becaus e mites and insects can
be carried into the laborator y on hair and clothing. Shower
and change clothes before entering the laborator y from the
field or greenhouse .
Determining the location of the tissue culture laborator y
is an importan t decision. Avoid locating it adjacen t to labora-
tories that handle microorganism s or insects or are used to
store seed s or other plant materials . Contaminatio n from air
vents and traffic can be a problem.
In designing a laborator y for tissue culture use, arrange
the work areas (media preparation , culture transfer , and
1