Strategies for improving the growth and development of crop species have been investigated for many years. Seed priming is a pre-sowing strategy for influencing seedling development by modulating pre-germination metabolic activity prior to emergence of the radicle and generally en¬hances germination rate and plant performance. During priming, seeds are partially hydrated so that pre-germinative metabolic ac¬tivities proceed, while radicle protrusion is prevented, then are dried back to the original moisture level. Common priming techniques include osmopriming, halopriming, hormopriming and hydropriming. Hydropriming and osmopriming contribute to significant improvement in seed germination and seedling growth in different plant species. Production of H2O2 during the early imbibitions period has been demonstrated; ROS produced after imbibition are assumed to play a role in seed germination. Thus, these reports suggest that ROS might play a signaling role in seed germination and dormancy. Although several lines of evidence indicate that ROS affect seed germination, there is little information establishing a direct link between the change in levels of ROS anti-oxidative activities and priming.

1. Hydropriming and Osmopriming Effects on Cowpea (Vigna
unguiculata L.): Involvement of oxidative stress
Presented by: BOUCELHA Lilya, ABROUS-BELBACHIR Ouzna, DJEBBAR Rèda *
Laboratory of Physiology and Organismal Biology, Faculty of Biological Sciences, University of Science and Technology Houari Boumediene
(USTHB), BP 39, El Alia, Bab Ezzouar, Algiers, Algeria corresponding author: reda_djebbar@yahoo.fr
INTRODUCTION
The International Congress of the German Botanical Society (Botanikertagung), 30 Sept-04 Oct 2013 in Tübingen, Germany
MATERIAL AND METHODS
Seed material:
Vigna
unguiculata
(Cowpea or bean of
black eye)
Origin: Argentina
Parts used: Seeds and
embryos
Control
No treatement
6 hours Imbibition
Imbibition 6 h in
distilled water
Kinetics of germination of Vigna unguiculata (cowpea) seeds
Hydropriming
Imbibition in distilled
water 3 h et 6 h +
redéshydration
Osmopriming
PEG 30%
6 hours Imbibition in
PEG 30%
RESULTS AND DISCUSSION
The biochemical parameters (in embryos)
Pretreatment of seeds
•The two forms of Glutathione (reduced and oxidized) were measured by spectrophotometer and by an enzymatic
method on microplates according Queval and Noctor (2007).
•Highlighting ROS is made by a cytological method: The hydrogen Peroxyde (H2O2) by 3,3’ diaminobenzidine (DAB)
and Superoxyde anion (O2
-) by Nitroblue tétrazolium (NBT).
Whatever the type of pretreatment hardening improves
germination performance.
The positive effects of the hardening that occur during
germination and emergence affect growth.
Thus, the effects of priming depends strongly on the duration
and type of treatment.
The hardening allows a more rapid and uniform germination,
better growth and a stronger tolerance to osmotic stress.
This can be the consequences of biochemical changes and
molecular level of the seed such as:
Modification of enzyme’s activities including those involved
in the degradation of reserves (alpha amylase, ...) and the
scavenging of ROS (SOD, APX, CAT, ...).
The activation of the endo-β-mannase which is the enzyme
responsible for the synthesis of ethylene (a hormone that
allows the degradation of the endosperm for dormancy)
The progress of cell cycle by synchronizing the replication
of DNA.
The synthesis of new proteins such as stress proteins under
genetic control by mRNA and DNA.
Stimulating the synthesis of osmolytes such as proline and
soluble sugars involved in osmotic adjustment in stressful
conditions.
(Gelormini, 1995; Gao and al., 1999; De Castro and al., 2000; Soeda
and al., 2004; Atia and al., 2006; Varier and al., 2010).
0
10
20
30
40
50
60
70
80
90
100
0 1 2 4 7
Temoin
Endurcie
Days
%ofgermination
Control
Primed
0,0
0,5
1,0
1,5
2,0
2,5
3,0
3,5
4,0
4,5
5,0
Control Primed
Pretreatment
Lengthsofrootsincm
Kinetics and germination capacity of seeds as pretreatments
The length of roots (after 5 days of setting
germination) and aerial parts of seedlings
(after 15 days) (in cm) from the control and
primed seeds
The
Germinatio
n
The
linear
Growth
In
condition
of osmotic
stress
(PEG 20%)
Glutathione,
reduced,
oxidized and
total
In embryo, the glutathione pool is present in appreciable amount in both forms (reduced and oxidized). It has been shown that the amount of total glutathione
decreased during germination (Klapheck, 1988; Kranner and Grill, 1993; Tommasi and al., 2001). This molecule is used in the biosynthesis of certain amino acids
involved in protein synthesis.
In our case, these hardened seeds were in rapid imbibition stage. Presumably, therefore, that during this phase, glutathione has been degraded into
aminoacids. This aminoacids were not incorporated into the protein as the protein content decreased in embryos hardened and amino acid content increased
(data not shown). Then the cell can recover nitrogen as glycine and glutamate, and sulfur in the form of cysteine (Penninckx, 2002).
Crop production and the establishment of good crops depend heavily on seed germination, which is a crucial step in the life cycle of higher plants (Cheng and Bradford, 1999). Yet germination may be
heterogeneous because the seeds do not germinate all the same way or at the same time. To solve these problems, called seeds undergo physiological conditioning appointed pre-germination or priming, which can
bring the seeds at the same physiological stage to synchronize germination and improve performance and stress tolerance (Heydecker and al., 1973; Heydecker, 1978; Taylor and al., 1998).
Priming is partially hydrated seeds at sufficient moisture level to allow pre-germination metabolic processes, but insufficient for the breakthrough of the radicle (during the reversible stage of germination)
(McDonald, 2000; Ghassemi -Golezani and al., 2010). Methods of seed priming can be divided into two groups according to the absorption of water is uncontrolled (hydro and hormopriming) or controlled (osmopriming)
(Taylor et al, 1998). It was clearly shown that the positive effects of this hardness were associated with various physiological, genetic, biochemical and molecular changes such as the mobilization of reserves, the
degradation of the endosperm, the activation of anti oxidative systems, the stimulation of the osmolytes synthesis, the activation of cell cycle and some genes involved in tolerance to abiotic stresses (Bray and al.,
1989; Dell'Aquila and Bewley, 1989; Davison and Bray, 1991; Bray, 1995; Castro and al., 2000; Vary and al., 2010).
Priming effect on the kinetics of seed germination in stressful conditions Priming effect on the kinetics of seed germination in stressful conditions
Control Osmoprimed
Embryons
The
cytological
detection of
ROS
Oxidized glutathione content in
control and primed embryos
The hydrogen
peroxide (H2O2)
The superoxide anion
(O2
-)
Control Hydroprimed
There is a large accumulation of O2
-, especially in the radicle. But in hardened embryos we notice
an accumulation of O2
- only at the ends of the radicle representing meristematic zones.
The DAB results show a weak accumulation of H2O2 in hardened embryos, but it is completely absent in
control embryos.
ROS, providing that their level of accumulation is tightly regulated by a balance between
production and elimination, appear to be beneficial for germination and, in particular, act as a positive
signal that can break dormancy of seeds hence the concept of “oxidative window for germination " (Bailly
and al., 2008). This new concept could explain the improved germinating performance in hardened seeds.
This explains the accumulation of ROS in certain tissues such as meristems (activation of cell division).
Thereby the interaction between ROS and hormone signaling pathways leads to changes in gene
expression or in cellular redox state.
CONCLUSION AND PERSPICTIVES
Seeds priming seem to be an effective and cheaper mean to improve germination, growth and even
stress tolerance of Vigna unguiculata.
Seeds priming causes physiological, biochemical and molecular changes highly regulated and
controlled by the expression of many genes.
Some consequences of this hardening are due to DNA methylation or the spatial conformation of
chromatin. Thus, epigenetic phenomena are of great importance for the understanding of many
phenomena in plant biology. They play a key role in the adaptation of plants to their environment.
Accordingly, the use of new molecular and genetic approaches are essential to better identifying the
expression of genes activated during priming.
Some REFERENCES
-Atia A., Debez A., Rabhi M., Habib-Urrehman-Athar H.U. and Abdelly C. (2006). Alleviation of salt-induced seed dormancy in the perennial halophyte
Crithmum maritimum L. (Apiaceae). Journal of Botany, 38 (5): 1367-1372.
- Bailly C ., El-Maarouf-Bouteau H. and Corbineau F. (2008). From intracellular signaling networks to cell death: the dual role of reactive oxygen species in seed
physiology. C.R. Biol., 331 (10) : 806-14.
-Bradford K. J. (1986). Manipulation of seed water relations via osmotic priming to improve germination under stress conditions. Hort Science, 21: 1105-1112.
- De Castro R. D., Van-Lammeren A. A. M., Groot S. P. C., Bino R. J. et Hilhors H. W. M. (2000). Cell division and subsequent radicle protrusion in tomato seeds
are inhibited by osmotic stress but DNA synthesis and formation of microtubular cytoskeleton are not. Plant Physiology., 122 : 327–335.
-Gao Y. P., Young L., Bonham-smith P. et Gusta L. V. (1999).Characterization and expression of plasma and tonoplast membrane aquaporins in primed seed of -
Brassica napus during germination under stress conditions. Plant Mol. Biol, 40 : 635-44.
-Gelormini G. (1995). Optimisation des propriétés germinatives des graines de colza par initialisation: aspects méthodologiques et fondamentaux. Thèse
nouveau doctorat, 171 p.
- Heydecker W. (1978). Primed seeds for better crop establishment. Span, 21 : 12-14.
-Varier A., Vari A. K. and Dadlani M. (2010). The subcellular basis of seed priming. Current Science, 99 (4) : 450-456.
0
10
20
30
40
50
60
70
80
90
100
0 1 2 3 4
Control
6h imbibition
3h hydro
6h hydro
PEG 30%
%ofgermination
Days
60
65
70
75
80
85
90
95
100
Control 6h
imbibi
3h
hydro
6h
hydro
PEG
30%
Control
6h imbibi
3h hydro
6h hydro
PEG 30%
Pretreatments
Capacitéofgermination(%)
Control 3h Hydropriming
6h HydroprimingPEG 30 %
2
4
6
8
10
12
Control 6h imbibi 3h hydro 6h hydro PEG 30%
Radicle
Aerial part
Priming
Lengthsincm
Control 3h Hydrprimingo 6h Hydropriming PEG 30 %
Control
Priming effect on the growth of plants grown from control seeds and hardened after 10 days of growth
7 Days
Control Osmoprimed
3h Hydropriming 6h Hydropriming
0
10
20
30
40
50
60
70
80
90
100
Témoins Endurcis
0
200
400
600
800
1000
1200
1400
1600
1800
Témoins Endurcis
Contentinmol/gFPM
Rate reduction of oxidized
glutathione in control and primed
embryos
Control Osmoprimed
Control Osmoprimed
%ofRatereduction
Hydroprimed
Control
Control