Under identical culture conditions, only 8 out

of 12 species and subspecies of the genus

Daucus proved

capable of somatic embryogenesis. Random amplified

polymorphic DNA analysis indicated a polymorphism

between the genomes of individual species that were capable

of embryogenesis and those that were not. Two

specific bands (1.1 kbp, 0.68 kbp) were detected only in

the genomes of individuals with the capacity for embryogenesis.

These were cloned and sequenced, and the homology

of the nucleotide sequences of the various species

was detected: this ranged from 74% to 92% for the

larger sequence and from 92% to 97% for the smaller

one. These DNA sequences would appear to be useful as

a marker of the capacity for somatic embryogenesis in

the genus



Daucus · DNA marker · Random amplified

polymorphic DNA · Somatic embryogenesis


IAA: Indole-3-acetic acid ·


Polymerase chain reaction · RAPD: Random

amplified polymorphic DNA ·


Two-dimensional polyacrylamide gel



Although there is a continuous steady increase in the

number of species of higher plants for which protocols to

induce somatic embryogenesis have been reported, little

is understood about this developmental process. Genetic

factors related to the recalcitrance to somatic embryogenesis

have been discussed for one or the other species and

even for varieties of one species (Vasil 1984), but no conclusive

evidence for a direct genetic correlation at the

molecular level has yet been reported. In order to gain a

better understanding of the significance of genetic factors

for detecting possible markers for somatic embryogenesis,

we conducted studies at the DNA level under identical

experimental conditions on selected regenerative and


species and subspecies.

In the investigation reported here we initially tested

the embryogenic capacity of 12

species or subspecies;

of these, 8 were successfully induced to undergo

somatic embryogenesis through the use of cultured petiole

explants. It appears that a finely tuned developmental

program is initiated in these explants when they are

placed into a medium containing an auxin. Following extensive

biochemical and histological investigations,

which also included a study of transgenic plant material

containing a mannopine synthase promoter/glucuronidase

reporter gene to observe auxin distribution within

cultured explants (Li and Neumann 1985; Neumann and

Grieb 1992; Neumann 1995, 2000; Grieb et al. 1997;

Imani 1999; Imani et al. 1999), we propose that this developmental

program is as follows.

Upon the initiation of embryogenesis, the endogenous

auxin is more or less evenly distributed among the cells

of the explanted petiole sections. After 3–4 days of culture

in an IAA-supplemented nutrient medium, a large

increase in cytoplasmic volume occurs near the vascular

bundles in cells that were previously vacuolated; this is

followed by the initiation of cell division. After 7 days of

culture, an accumulation of IAA can be observed in cells

near the vascular bundles, leaving the other tissue almost

free of IAA. Concurrently, the initiation of root primor-

Communicated by K.K. Kamo

J. Imani · L. Tran Thi · B. Arnholdt-Schmitt · C. Lein

K.-H. Neumann (


Institute of Plant Nutrition, Division of Plant Tissue Culture,

Justus Liebig University, Heinrich-Buff-Ring 26-32,

35392 Giessen, Germany

e-mail: Karl-Hermann.Neumann@ernaehrung.uni-giessen.de

Tel.: +49-641-9939190, Fax: +49-641-9939199

G. Langen

Institute for Phytopathology, and Applied Zoology,

Justus Liebig University, Heinrich-Buff-Ring 26-32,

35392 Giessen, Germany

S. Roy · A. Kumar

Biotechnology Laboratory, Department of Botany,

University of Rajasthan, Jaipur 302004, India

Plant Cell Rep (2001) 20:537–541

DOI 10.1007/s002990100363


J. Imani · L. Tran Thi · G. Langen

B. Arnholdt-Schmitt · S. Roy · C. Lein · A. Kumar

K.-H. Neumann

Somatic embryogenesis and DNA organization of genomes

from selected

Daucus species

Received: 17 November 2000 / Accepted: 22 May 2001 / Published online: 5 July 2001

© Springer-Verlag 2001