Plant Molecular Biology 43: 537–544, 2000.
© 2000 Kluwer Academic Publishers. Printed in the Netherlands.
A transgenic rice cell lineage expressing the oat arginine decarboxylase
(adc) cDNA constitutively accumulates putrescine in callus and seeds but
not in vegetative tissues
, Ludovic Bassie
, Olivia Lepri, Isaac Kurek
, Paul Christou and Teresa
Molecular Biotechnology Unit, John Innes Centre, Norwich Research Park, Colney Lane, Norwich, NR4 7UH, UK
author for correspondence;e-mail: firstname.lastname@example.org);
Present address: Department of Botany,
Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel;
these authors contributed equally to this work
Received 19 November 1999; accepted in revised form 28 April 2000
Key words: arginine decarboxylase, maize ubiquitin 1 promoter, Oryza sativa, polyamines.
We introduced the oat adc cDNA into rice under the control of the constitutive maize ubiquitin 1 promoter.
We studied molecularly and biochemically sixteen independent transgenic plant lines. Signiﬁcant increases in
mRNA levels, ADC enzyme activity and polyamines were measured in transgenic callus. These increases were not
maintained in vegetative tissue or seeds in regenerated plants, with the exception of one lineage. This particular
lineage showed very signiﬁcant increases in putrescine preferentially in seeds (up to 10 times compared to wild
type and controls transformed with the hpt selectable marker alone). We have demonstrated that in cereals such
as rice, over-expression of the oat adc cDNA results in increased accumulation of polyamines at different stages
of development. We have also demonstrated that strong constitutive promoters, such as the maize ubiquitin 1
promoter, are sufﬁcient to facilitate heritable high-level polyamine accumulation in seed. Our results demonstrate
that by screening adequate numbers of independently derived transgenic plants, it is possible to identify those
individuals which express a desired phenotype or genotype.
Polyamines (PAs) are a group of low-molecular-
weight polycationic compounds that are ubiquitous
in nature and important for many cellular processes.
In animals, PAs are known to play important roles
in DNA replication, transcription and protein syn-
thesis, and are therefore vitally important for cell
division, growth and development. All dividing cells
can synthesize PAs. The simplest PA, putrescine,
is derived from ornithine by ornithine decarboxy-
lase (ODC). Putrescine may then be converted into
the longer aliphatic PAs spermidine and spermine
by spermidine and spermine synthase, respectively,
which add propylamino groups generated from S-
adenosylmethionine (SAM) by SAM decarboxylase
(SAMDC; Malberg et al., 1998). In plants, putrescine
may be synthesized from either ornithine or arginine,
the former through the ODC pathway as found in
animals, and the latter through the alternative argi-
nine decarboxylase (ADC) pathway, involving two
intermediates, agmatine and N-carbamoylputrescine
(Malmberg et al., 1998).
Food varies widely in its PA content and distri-
bution, with mature Cheddar cheese, for example,
containing very high levels, particularly putrescine,
and cerealgrains containing low levelsof all PAs (Bar-
docz et al., 1993). Cereal grain represents the staple
diet for most people in the world, yet is deﬁcient in
many essential nutrients. Rice is probably the most
importantcereal, accountingfor most of the energy in-
takefor up to 50% of the world’s population(Christou,
1994). However, rice containsvery low levels of nutri-
ents such as iron, calcium and PAs (Bardocz et al.,