Background and aim: Pre-germination treatments represent the physiological methods that improve plant production by modulating the metabolic activities of germination before the emergence of the radicle. This treatment consists of an osmotic (osmopriming) or a hormonal (hormopriming) and/or redehydration (hydropriming) treatments. The approach allows the elimination of dormancy, homogenization (synchronization) of germination, better growth, earlier flowering and a tolerance to abiotic stresses such as drought and salinity. It was suggested that ROS play a crucial role in signaling seed germination. Our work consisted in studying changes in the redox status in the embryonic axis (radicle and plumula ) and in cotyledons without and with priming, by estimating antioxidant activities and production of reactive oxygen species (ROS). Methods: In the cotyledons, radicle and plumula taken from control Vigna unguiculata seeds and having undergone osmopriming, simple hydropriming and double hydropriming some antioxidant enzymatic activities as well as total non-enzymatic antioxidant capacity (TAC) were measured. The production of hydrogen peroxide (H2O2) by DAB and superoxide anion (O2-) by NBT was also assessed. Results: The results obtained showed that priming allows activation of antioxidant enzymes, especially in the plumula. On the other hand, our results showed that priming caused an accumulation of ROS in embryonic tissues. This may also explains the improvement of seed germination performance according to the oxidative window model. Conclusion: Priming induced changes in the redox environment at the seed level. These changes were closely related to the type of seed initiation. Indeed, a double cycle of hydration-rehydration induced the broadest spectrum of modifications of the redox status by inducing better germination and better growth under favorable or unfavorable conditions.

1. Vigna unguiculata (L.) Walp. seed priming is related to redox status
of plumule, radicle and cotyledons.
Presented by: Lilya BOUCELHA, Ouzna Belbachir-Abrous and Réda DJEBBAR
Laboratory of Biology and Physiology et of Organisms, Faculty of Biological Sciences, University of Sciences and Technologie Houari
Boumediene (USTHB), BP 39, El Alia, Bab Ezzouar, Algiers –Algeria
liliaboucelha@yahoo.fr /oabrous@yahoo.fr/ reda_djebbar@yahoo.fr
INTRODUCTION
The International Days of Biotechnology (IDB 2018) 20 - 23 December 2018 Sousse (Tunisia)
MATERIEL AND METHODES
Témoin
Any traitement
before germination
6h imbibition
Soaking for 6 hours
in distilled water
before germination
Simple Hydropriming
Soaking for 3 hours or
6 hours in distilled water
+ Dehydration
Osmopriming
PEG 30 %
Soaking for 6 hours
in the PEG at 30%
RESULTS AND DISCUSSION
Priming of seed
CONCLUSION AND PERSPECTIVES
REFERENCES BIBLIOGRAPHIQUES
-Bailly C., El-Maarouf-Bouteau H., Corbineau F. (2008). From
intracellular signaling networks to cell death: the dual role of reactive oxygen
species in seed physiology. Comptes Rendus Biologies 331, 806-814.
-Beckers GJM., Jaskiewicz M., Liu Y., Underwood WR., He SY., Zhang
-Boucelha L and Djebbar R. (2015). Influence de différents traitements
de prégermination des graines de Vigna unguiculata (L.) Walp. sur les
performances germinatives et la tolérance au stress hydrique. Biotechnol.
Agron. Soc. Environ., 19(2): 132-144.
-Boucelha L. (2015). Compréhension des mécanismes régissant
l’endurcissement des graines de Vigna unguiculata (L.) Walp. Thèse de
doctorat, USTHB 195 p
-Boucelha L, Djebbar R and Abrous-Belbachir O (2019). Vigna unguiculata
(L.) Walp. seed priming is related to redox status of plumule, radicle and
cotyledons. Functional Plant Biology DOI 10.1071/FP18202
-Bradford K.J. (1986). Manipulation of seed water relations via osmotic
priming to improve germination under stress conditions. Hort Science., 21:
1105-1112.
-Heydecker W., Higgins J., Gulliver R.L. (1973). Accelerated
germination by osmotic seed treatment. Nature 246, 42-44.
-Varier A., Vari A.K. and Dadlani M. (2010). The subcellular basis of
seed priming. The anthors are in the Indian Agricultural Research Institute,
Current Science., 99.
- Yang L., Zhang J., He J., Qin Y., Hua D., Duan Y., Chen Z. and Gong
Z. (2014). ABA-Mediated ROS in mitochondria regulate root meristem
activity by controlling PLETHORA Expression in Arabidopsis. Plos Genetics
Journal., 10, e1004791.
Pre-germination treatments represent the physiological methods that improve plant production by modulating the metabolic activities of germination before the emergence of the
radicle. This treatment consists of an osmotic (osmopriming) or a hormonal (hormopriming) and/or redehydration (hydropriming) treatments. The approach allows the elimination of
dormancy, homogenization (synchronization) of germination, better growth, earlier flowering and a tolerance to abiotic stresses such as drought and salinity. Seed priming causes
biochemical, physiological, molecular, and genetic changes such as the accumulation of latent signaling components that will be used during a re-exposure to a stress (Heydecker et al.
1973; Bradford 1986; Beckers et al. 2009; Boucelha and Djebbar 2015).
The concept of oxidative window for germination proposed by Bailly et al. (2008) could also explain the improvement of germination performance of primed seeds. According to this
theory, oxidative stress was essential for triggering germination. ROS were produced continuously during seed development, from embryogenesis to germination. They play a dual role in
the physiology of seeds by being involved in the pathways of cell signaling on one hand and acting as toxic products accumulated under stress conditions, on the other hand.
Vegetal Materiel:
Vigna unguiculata
(Black-eyed bean)
Origin: Madagascar
Used Parts: Seed and
embryos (cotyledons,
radicle and plumule)
Double
Hydropriming
Double cycle of soaking
3 hours in distilled water
followed by Dehydration
Germination
Speed
and
Capacity
Effects of different priming methods on the
germination of Vigna unguiculata seeds in
distilled water.
Catalase
Activity
Superoxide
Dismutase
activity
Priming, and especially
double dehydration,
improved seed germination
performance of Vigna
unguiculata. Boucelha and
Djebbar (2015) has shown
the beneficial effects of
different types of priming
on the germination and
growth of Vigna unguiculata
plants under favorable and
stressful conditions.
Seed priming have stimulated the
production of ROS in the embryonic
tissues and essentially in the meristematic
zones, which reflected the strong
activation of cell division during priming.
Indeed, Yang et al. (2014) suggested that
an accumulation of ROS in meristematic
cells played a major role in signaling root
growth. These results suggested that the
beneficial effects of priming on improved
germination performance could be
explained by an oxidative window as
proposed by Bailly et al. (2008).
Cotyledons
Radicle Plumule
1. Detection of hydrogene peroxide (H2O2)
3,3’ diaminobenzidine
(DAB)
Polymerization with
H2O2 Oxidoreduction
Very stable
Brown
precipitate
2. Detection of superoxide anion (O2
-)
Tetrazolium
Nitroblue (NBT)
Reduction by
O2
-
Stable blue-
indigo
Blue formazan
Cytochemical analyzes of ROS
Photographs showing the detection of hydrogen peroxide (H2O2) by DAB in embryos from the control and
treated seeds (A) and a representative embryo of each batch taken under a binocular magnifying glass with
8x magnification (B).
Photographs showing the detection of the superoxide anion (O2
-) by NBT in embryos from the control and
treated seeds (A) and a representative embryo of each batch taken under a binocular magnifying glass with 8x
magnification (B).
0
50
100
150
200
250
300
350
400
Cotyledons Radicle Plumule
SuperoxidedismutaseActivity
(UnitsofSOD.min-1mg-1ofprot)
Control 6h imbib PEG 30% 3h hydro 6h hydro 3h double hydro
Superoxide dismutase activity in cotyledons, radicle and
plumule from the control and treated Vigna unguiculata
seeds.
0
1
2
3
4
5
6
Cotyledons Radicle Plumule
Control 6h imbib PEG 30% 3h hydro 6h hydro 3h Double hydro
CatalaseActivity
(µmolH2O2mn-1mg-1prot)
Hydrogene
peroxide
Production
Catalase activity in cotyledons, radicle and plumule from the
control and treated Vigna unguiculata seeds.
Anion
Superoxide
Production
Pre-germination treatments stimulated
the activation of antioxidant enzymes in
the cotyledons and the embryonic axis.
This activation depended closely on the
type of treatment and even on the organ,
which was related to the best germination
performance and tolerance improvement
to osmotic stress (Boucelha and Djebbar
2015). Activation of these enzymes was
probably triggered in response to seed
dehydration during the seed priming.
Control PEG 30%
6h hydro 3h Double hydro
0
20
40
60
80
100
0 1 2 3 4 5
Control
6h imbib
3h hydro
6h hydro
3h Double
hydro
PEG 30%
Pourcentageof
germination
Days
0
20
40
60
80
100
120
Control 6h imbib PEG 30% 3h hydro 6h hydro 3h double
hydro
Germination
capacity(%)
+918 %
+493 %
+25 %
+256 %
+96 %
-40 %
+871 %
a a
b
c
d e
a
b
e
f
c
d a1 a2
b1
b2c1
c2
d1
e1
f1
d2
e2
f2
a
b
d
b
c
d
a1
b1
c1
b1
b1
d1
a2
b2
c2
d2
d2
e2