Table Of ContentHeidelberg
Science
Library
Plasmids of Eukaryotes
Fundamentals und Applications
By K. Esser
U. Klick C. Lang-Hinrichs P. Lemke
H. D. Osiewacz U. Stahl P. Tudzynski
With 25 Figures
Springer-Verlag
Berlin Heidelberg New York Tokyo
ISBN-13: 978-3-540-15798-4 e-ISBN-13: 978-3-642-82585-9
DOl: 10.1007/978-3-642-82585-9
Library of Congress Cataloging-in-Publication Data. Main entry under title: Plasmids of
eukaryotes. (Heidelberg science library) Bibliography: p. Includes index. 1. Plasmids.
2. Eukaryotic cells. 3. Genetic engineering. I. Esser, Karl, 1924-. II. Series.
QH452.6.P57 1986 574.87'328 85-27749
This work is subject to copyright. All rights are reserved, whether the whole or part of the
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© by Springer-Verlag Berlin Heidelberg 1986
The use of registered names, trademarks, etc. in this publication does not imply, even in the
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2131/3130-543210
Dedicated to
GEORGES RIZET
the discoverer of
Podospora anserina
Preface
The possession of plasmids was for a long time recognized only in the
bacteria. It is now evident that plasmids, or replicative forms of DNA
structurally and experimentally comparable to bacterial plasmids, exist
in eukaryotic organisms as well. Such plasmids are in fact common
among fungi and higher plants. The present review is undertaken to
provide a comprehensive account of the data available on plasmids
found in eukaryotic organisms. This review will not consider plasmids
of prokaryotic origin, even though certain bacterial plasmids, such as
the tumor-inducing (Ti) plasmids of Agrobacterium tumefaciens, may
be intimately associated with transformation of the eukaryotic host.
This book, moreover, does not consider transformation experiments in
eukaryotic hosts involving viral DNA as vectors, although indeed such
vectors have been developed for use in plant and animal systems.
After a general introduction, providing historical perspective on the
nature and role of plasmids, a list of eukaryotic plasmids will be
presented according to their origin. This is followed by a detailed
discussion of known structure and function. In subsequent chapters the
practical implications of eukaryotic plasmids for molecular cloning and
biotechnology will be discussed. This latter part traces the development
of interest'in biotechnical genetics and gives special consideration to the
use of eukaryotic systems for gene cloning. The terminology biotechni
cal genetics is introduced to the reader and is used in a general sense as
equivalent to genetic engineering. Biotechnical genetics includes, but is
not limited to, gene cloning through recombinant DNA technology.
Genetic manipUlations involving protoplast fusion, embryo transplanta
tion or directed mutagenesis would also represent forms of biotechnical
genetics.
Since this booklet is intended as a general reference source, not only
for scholars but for industrial scientists and engineers as well as others
more generally interested in biotechnology, a concerted effort has been
made to compile recently published scientific data along with relevant
background information and experimental details. The authors invite
constructive criticism from readers of this first edition concerning the
selection and presentation of material in the text.
During the preparation of the manuscript, friends and colleagues
have assisted with critical advice. We would like to acknowledge also
the assistance of Frau Ch. Konig and Frau D. Lenke in preparing the
manuscript and of Herr H. J. Rathke for the excellent art work.
Bochum, June 1985 The Authors
Contents
I. Introduction. 1
A. Definition. . 1
B. Historical Perspective 2
II. Fundamental Aspects 7
A. General Characteristics 7
B. Nuclear Plasmids ... 13
1. Saccharomyces cerevisiae - the 2,um Plasmid 13
2. Dictyostelium discoideum - a Cobalt Resistance Plasmid 21
3. Drosophila melanogaster - the Transposable Element
Copia ....... 23
C. Mitochondrial Plasmids 27
1. Podospora anserina - the Senescence Plasmid 27
2. Neurospora crassa - the Stopper and Poky Plasmids 34
3. Neurospora crassa - the Mauriceville Plasmid;
Neurospora intermedia-the Labelle and Fiji Plasmids 37
4. Claviceps purpurea 41
5. Other Fungi . . . . 44
6. Higher Plants . . . . 48
D. Unknown Association . 53
III. Practical Implications 57
A. Fundamentals for Eukaryotic Gene Cloning 58
1. Generalized Vector ....... . 59
2. Choice of an Appropriate Host Cell 64
B. Plasmids for Gene Cloning. . . . . . . 65
1. The 2,um Plasmid of Saccharomyces cerevisiae 65
2. RibosomalDNAPlasmids . . . . . . . . . . 68
3. The Senescence Plasmid of Podospora anserina . 69
4. The Labelle Plasmid of Neurospora intermedia 69
5. The Mitochondrial Plasmid of Cochliobolus heterostro-
phus . . . . . . . . . . . . . . . . . . . . . . . . . . 70
6. The Transposable Elements of Drosophila melanogaster 70
C. Organellar DNA for Gene Cloning ............. 71
1. Vectors Based on Confirmed Replication Origins . . . .. 73
2. Vectors Based on Random DNA Segments as Origin of
Replication. . . . . . . . . . . . . . . 74
D. Maintenance of Vector Transferred Genes . 79
1. Stabilization of Vectors in Host Cells . 79
2. Efficient Expression of Cloned Genes 80
E. Biotechnological Perspectives 87
References . . 89
Subject Index . 119
Authors
Prof. Dr. Dr. h. c. KARL ESSER
Lehrstuhl fur Allgemeine Botanik, Ruhr-UniversiHit, Postfach 102148,
D-4630 Bochum
Dr. ULRICH KOCK
Lehrstuhl fur Allgemeine Botanik, Ruhr-Universitat, Postfach 102148,
D-4630 Bochum
Dr. CHRISTINE LANG-HINRICHS
Technische Universitat Berlin, Pachgebiet Mikrobiologie, SeestraBe 13,
D-1000 Berlin 65
Prof. Dr. PAUL LEMKE
Molecular Genetics Program, Department of Botany and Microbiology,
Auburn University, Alabama 36849, USA
Dr. HEINZ DIETER OSIEWACZ
Lehrstuhl fur Allgemeine Botanik, Ruhr-Universitat, Postfach 102148,
D-4630 Bochum
Prof. Dr. ULF STAHL
Technische Universitat Berlin, Pachgebiet Mikrobiologie, SeestraBe 13,
D-1000 Berlin 65
Priv. Doz. Dr. PAUL TUOZYNSKI
Lehrstuhl fur Allgemeine Botanik, Ruhr-Universitat, Postfach 102148,
D-4630 Bochum
I. Introduction
Concurrent with the development of bacterial genetics in the early
1950s was the discovery in Escherichia coli of genetic factors not lo
calized routinely on the bacterial chromosome. These included:
1. The fertility (F +) factors responsible for bacterial conjugation;
2. Factors responsible for the production of the bacterial toxins of
the colicin type;
3. Factors responsible for bacterial resistance to antibiotics. It be
came evident in time that these factors, termed plasmids by
Lederberg (1952), consisted of double-stranded DNA (dsDNA)
and were able to propagate in either of two alternative modes:
either autonomously in the bacterial cytoplasm (replicative plas
mids) or as an integral part of the bacterial chromosome (inte
grative plasmids).
Before these dual modes of plasmid replication were understood, inserted factors
were called episomes and considered to be fundamentally different from the extra
chromosomal plasmids (see Bresch 1964).
The autonomous or extrachromosomal mode of replication explains
why bacterial cells may contain many copies of specific plasmids. Plas
mids are found in a great variety of bacteria and vary considerably in
size, ranging from 2.25kb to 500kb.
A. Definition
In short, one may define a plasmid as any genetic element which is
supplemental to the normal genome of the cell (modified after Rieger et
al. 1976). Details and further references may be taken from textbooks,
such as Bukhari et al. (1977); Knippers (1982); Fincham (1983) and
Kaudewitz (1983). Thus broadly defined, a plasmid may be either
extragenomic (exoplasmid) or derived from· the cell's normal genome
as a sequence brought to multicopy status by autonomous replication
(endoplasmid). In this context it is necessary to discuss terminology
related to plasmid replication.
As early as 1963, Jacob and Brenner introduced the term repli
con for the smallest unit of autonomous replication according to data
obtained in prokaryotes. Subsequently, it was found that this unit
starts with a specific sequence responsible for initiations of replica
tion and ends with a terminator sequence. It was generally accepted
to call the initiation sequence the origin of replication: its function
consists in formation of the replication fork, visible either by electron
microscopy or by fiber autoradiography (Kornberg 1980). Sometimes
a proper identification of the origin of replication was not possible
either due to a lack of material or to problems in technology. In these
cases DNA sequence analysis can provide indicative evidence for its
location. Then, the term putative origin of replication is used.
Another technique to identify a putative origin of replication has
been developed in yeast (p. 74 f.). By shot gun cloning experiments
DNA sequences can be identified as putative origins after integration
in nonreplicative vectors (Fig. 24) and are termed ars (autonomously
replicating sequences). From more developed analysis it has become
obvious that .some ars, although functional in yeast, fail to replicate
in other eukaryotic systems. Therefore, the term ars is restricted to
sequences tested in yeast. For reasons of simplification and to avoid
confusion for our reader, in this booklet all sequences promoting au
tonomous replication are called origins of replication (on), regardless
of whether they function in yeast or in other systems. While struc
turally the majority of known plasmids are covalently closed and cir
cular double-stranded DNA (cee dsDNA) molecules, linear dsDNA
plasmids are also recognized. Self-replicating forms of single-stranded
RNA (ssRNA), such as the viroids of higher plants, and endogenous
dsRNA molecules found in certain fungi might also be included in a
broad conceptual definition of a plasmid. Since reviews are available
on viroids (Diener 1984) and dsRNA plasmids of fungi (Tipper and
Bostian 1984), these genetic elements will not be considered in the
present review.
B. Historical Perspective
Aside from the role played by plasmids in bacterial recombination,
early interest in bacterial plasmids was focused during the 1960s on
antibiotic resistance and the practical implications for control of ac
quired resistance. It was found that eoliplasmids were able to infect
other bacteria, such as Salmonella typhosa, and were expressed in
the new host, leading to interspecific transfer of antibiotic resistance.
It also became clear that plasmids were not restricted to E. coli but
rather common among bacteria (for rev. see Foster 1983). The field
of plasmid research received new emphasis in the 1970s through the
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