Glycine betaine application modifies biochemical attributes of osmotic adjustment in drought stressed wheat

ORIGINAL PAPER

Glycine betaine application modifies biochemical attributesof osmotic adjustment in drought stressed wheat

Neha Gupta • Sanjeev Kaur Thind •

Navtej Singh Bains

Received: 14 February 2013 / Accepted: 3 October 2013 / Published online: 10 October 2013

� Springer Science+Business Media Dordrecht 2013

Abstract Nineteen wheat genotypes were used to

examine the effects of foliar applied glycine betaine (GB,

100 mM) on concentration of various osmolytes (such as

proline, choline, GB and sucrose) under drought stress

conditions. Drought stress caused a significant increase in

proline content and GB content of wheat genotypes, both at

maximum tillering and anthesis stages. Choline and

sucrose were accumulated significantly at higher levels

under stress conditions at both the stages. GB application

increased the proline content and endogenous levels of GB

in comparison to their stressed counterparts both at maxi-

mum tillering and anthesis stages but this increase was

observed to be genotype specific. Furthermore, significant

decrease in choline levels and sucrose contents of GB

treated plants at anthesis stage and enhanced levels of

proline questioned about involvement of GB in production

of other osmolytes as well as stage specific response of

wheat genotypes to GB spray. But these changes in

osmolyte accumulation (OA) were not correlated with

relative water content and stress tolerance index observed,

under both GB sprayed and non-sprayed drought stressed

conditions. So OA could not be considered as a selection

criteria for drought tolerance in wheat.

Keywords Drought stress � Wheat � Proline �Glycine betaine levels

Abbreviations

GB Glycine betaine

OA Osmolyte accumulation

RWC Relative water content

STI Stress tolerance index

Introduction

Wheat is a cereal of choice and a staple food for people

all over the world. About 230 million ha of land is used

for wheat cultivation worldwide and half of this area is

routinely afflicted with drought stress (Trethowan and

Reynolds 2007). As one of the abiotic constraints, drought

determines yield and quality of wheat, through sink

strength and source capacity. Losses in grain yield due to

drought may range from 17 to 70 %. Drought stress

involves not only water deprivation but also nutrient

limitations, salinity and oxidative stress to plants. Among

the various protection mechanisms of plants, accumula-

tion of osmolytes/compatible organic solutes is important

for ensuring survival under stress conditions. These

compatible solutes are of low molecular weight, having

high solubility in water and are non-toxic even at high

concentrations (Bohnert et al. 1995). Osmolyte accumu-

lation (OA), usually called osmotic adjustment, has long

been proposed as a selection criterion in traditional

breeding or through marker assisted selection and genetic

engineering programmes to generate drought tolerant

crops (Ludlow and Muchow 1990; Zhang et al. 1999). A

wide variety of osmolytes including betaines, polyols,

sugars and amino acids etc. exist in the plant kingdom

(Rhodes and Hanson 1993). Among the many betaines

N. Gupta (&) � S. K. Thind

Department of Botany, Punjab Agricultural University,

Ludhiana 141004, India

e-mail: [email protected]

N. S. Bains

Department of Plant Breeding and Genetics, Punjab Agricultural

University, Ludhiana 141004, India

123

Plant Growth Regul (2014) 72:221–228

DOI 10.1007/s10725-013-9853-0

and its related compounds known in plants, glycine

betaine (GB) occurs most abundantly in response to

dehydration stress (Mohanty et al. 2002; Yang et al.

2003). GB (N,N0,N00-trimethylglycine) is a quaternary

ammonium compound that is naturally synthesized by a

large number of organisms, including members of plant

families Chenopodiaceae, Amaranthaceae, Graminae,

Compositae and Malvaceae (Rhodes and Hanson 1993;

Gorham 1996; Blunden et al. 1999). In addition to its

function in osmoregulation, GB plays an important role in

stabilization of complex proteins, enzymes (e.g., Rubisco)

and membranes (Bohnert and Jensen 1996; Allakhverdiev

et al. 2003), besides protection of photosynthetic appa-

ratus from photodamage (Sakamoto and Murata 2002;

Zhao et al. 2007) and maintenance of electron flow in

thylakoid membranes (Ma et al. 2006) under various

stress conditions. Moreover, involvement of GB in regu-

lation of the activities of antioxidant enzymes has been

reported (Ma et al. 2006; Park et al. 2007; Raza et al.

2007; Wang et al. 2010).

Transgenics for enzymes involved in GB synthesis

[such as betaine aldehyde dehydrogenase (BADH), cho-

line dehydrogenase (CDH), choline monooxygenase

(CMO) and choline oxidase (COD)] in rice (Sakamoto

et al. 1998), tomato (Park et al. 2007), Arabidopsis (Alia

et al. 1999), tobacco (Ma et al. 2007) and wheat (Wang

et al. 2010) exhibited an enhancement in tolerance to

cold, drought, heat and salinity stress. Inability to pro-

duce sufficient amounts of GB in some of these trans-

genics has been attributed to low availability of its

substrate choline, reduced transport of choline across the

chloroplast envelope and the high energy cost of GB

biosynthesis. In this context, an alternative approach

involving exogenous application of GB has gained

attention. There are many reports demonstrating positive

effects of exogenous application of GB on growth, sur-

vival and tolerance of a wide variety of GB accumulator/

non accumulator plants under various stress conditions

(Rajasekaran et al. 1997; Diaz-Zorita et al. 2001). Foliar

application of GB in wheat was also found to alleviate

photoinhibition (Ma et al. 2006), improve photosynthesis

via altering lipid composition of thylakoid membranes

(Zhao et al. 2007) and maintain higher antioxidant

enzyme activities under drought and salinity stress (Raza

et al. 2007; Liang et al. 2009). But none of these studies

were carried out under long term field stress conditions

where multiple stresses and complexity in stress response

occurs. Moreover, little information was available on

exogenously applied GB affecting other osmolytes levels

in wheat genotypes differing in drought tolerance.

Keeping in view the above points, the present study

aimed at understanding the role of applied GB in terms

of changes induced in levels of various compatible

osmolytes and their contribution to water deficit tolerance

in wheat under field conditions.

Materials and methods

Seeds of eight cultivars (C306, PBW175, PBW527,

PBW343, PBW550, DBW17, PBW 621 and HD 2967) and

eleven advanced lines (BW9022, BW9097, BW9183,

BW9005, BW9016, BW8989, BW8366, BW9151,

BW8362, BW9025 and BWL0089) of wheat were obtained

from Department of Plant Breeding and Genetics, Punjab

Agricultural University, Ludhiana. Field experiments were

conducted during the growing seasons of 2010–2011 and

2011–2012. The experiment was laid out in a split plot

design with three main plot treatments (i.e. control or

normal irrigated, drought stress and drought stress with GB

application) and 19 wheat varieties as subplot treatments.

Each main plot treatment was tested by three replications

with a subplot size of 2 9 3 m2. Wheat genotypes were

sown in field according to standard agronomic practices at

a seed rate of 40 kg/acre and row to row spacing of 23 cm.

Drought stress was imposed by withholding irrigation

throughout the season, except for a pre-sowing irrigation to

ensure optimum germination. Drought stressed plots were

covered with plastic sheets during rains. Control plots were

provided with five irrigations (each of 8–10 cm depth)

during the crop life cycle. In case of GB sprayed plots,

aqueous 100 mM GB solution containing 0.1 % TWEEN

20 was sprayed on the leaves until run off twice a day for

2 days as described by Ma et al. (2006) at maximum til-

lering and anthesis stages. The two types of GB non-

sprayed plots i.e. control (normal irrigated) and drought

stressed plots were sprayed with only water containing

0.1 % TWEEN 20 at both the stages. After 1 week of GB

application, leaf samples were collected from each plot.

The youngest fully expanded leaf and flag leaf from 10

random plants were excised for biochemical estimations at

maximum tillering and anthesis stages respectively.

Determination of free proline

The proline content was determined spectrophotometeri-

cally by the ninhydrin method as described by Bates et al.

(1973). Leaf samples (0.250 g) from each replicate were

homogenised in 5 ml of 3 % sulphosalicylic acid for pro-

line extraction. The filtrate (2 ml) was then reacted with

acid ninhydrin and glacial acetic acid at 100 �C for 1 h.

After termination of the reaction in an ice bath, extraction

was done using toluene and absorbance of toluene con-

taining the pink coloured chromophore was determined at

520 nm. The proline concentration was determined from a

standard curve and calculated on fresh weight basis.

222 Plant Growth Regul (2014) 72:221–228

123

Determination of glycine betaine and choline content

Leaf GB and choline was determined following Grieve and

Grattan (1983). Extraction of GB and choline was done

from dried leaf material (0.5 g) in deionized water (20 ml)

for 24 h at 25 �C. The extract (0.5 ml) was then mixed

with 0.5 ml of 2 N H2SO4 and cooled in an ice-bath for an

hour. Then after addition of 0.2 ml potassium tri-iodide

solution, the contents were gently mixed and placed in an

ice-bath for 16 h. GB periodide crystals obtained were then

dissolved in 9 ml of 1,2-dichloroethane and optical density

of the solution was measured at 365 nm.

For choline estimation, the aqueous extract (0.5 ml)

obtained from dried leaf material was mixed with 0.5 ml of

0.2 M potassium phosphate buffer (pH 6.8) instead of 2 N

H2SO4. The other steps followed were same as for GB

estimation. The concentrations of choline and GB were

calculated on the dry weight basis.

Determination of sucrose

Sucrose content was estimated using the resorcinol method

of Roe et al. (1949) and extraction of sugars was done by

ethyl alcohol.

Relative leaf water content and stress tolerance index

(STI)

Leaf relative water content (RWC) was calculated via leaf

discs by the formula (FW-DW/TW-DW) 9 100 %, where

FW was fresh weight, TW was the turgid or saturated fresh

weight and DW was the dry weight after oven drying of leaf

discs at 80 �C for 48 h (Ma et al. 2007). The STI was cal-

culated by the formula: Yd 9 Yp/( �Yp)2 where Yd was mean

grain yield of a genotype under drought, Yp was mean grain

yield of a genotype under control conditions and �Yp was

mean grain yield of all genotypes under control conditions.

Statistical analysis

All measurements were recorded on three replications. The

data were subjected to statistical analysis by means of factorial

experiment in a random block design using two way ANOVA,

consisting of 19 genotypes and treatments at 3 levels.

Results

Proline content in wheat leaves

Free proline content in leaves was significantly (P B 0.05)

higher in drought stressed plots (pooled across genotypes)

as compared to irrigated plots (Fig. 1a, b). Further, this

increase was more pronounced at maximum tillering stage

(181.01 %) than at anthesis (96.09 %). Not all genotypes,

however, followed this trend and four exceptions were

observed. Application of GB to drought stressed plots

enhanced proline accumulation further both at maximum

tillering (114.7 %) and anthesis stages (147.5 %). Inter-

estingly, the level of enhancement of proline content as a

result of GB application was seemed to depend upon the

inherent levels prior to spray. Overall nine genotypes

(C306, PBW 175, BW 9097, BW 9183, BW 8989, BW

9151, BW 8362, BWL0089 and PBW 621) at maximum

tillering stage and fourteen genotypes at anthesis stage

(C306, PBW 527, PBW 343, DBW 17, BW 9022, BW

9097, BW 9183, BW 9005, BW 8989, BW 8366, BW

8362, BWL0089, PBW 621 and HD2967) showed this

response in proline accumulation with GB spray under

drought stress conditions. Most of the genotypes showing

relatively lower proline levels prior to spray exhibited a

larger magnitude of increase on GB application. Genotype

specific response was also reflected in seven genotypes

(PBW175, PBW 343, BW 9183, BW 8989, BW 9151, BW

8362 and BWL0089) wherein GB application changed the

stage response (maximum tillering versus anthesis) at

which higher proline levels were observed.

Choline and glycine betaine contents

As depicted in Fig. 2a and b, choline content of drought

stressed plots showed a significant increase over irrigated

ones. The increase was of a much higher magnitude at

maximum tillering stage (111.5 %) in comparison to

anthesis stage (12.2 %). The actual content of choline,

however was lower at maximum tillering stage. Contrast-

ing effect of GB spray on choline levels was observed at

the two stages. While GB application enhanced choline

levels at maximum tillering stage (44.5 %), significant

reduction (-70.4 %) was observed at anthesis stage.

Importantly, this reduction in choline content was consis-

tent across genotypes.

In case of GB content, drought stressed plots had sig-

nificantly higher levels of GB in comparison to irrigated

plots (Fig. 3a, b). This enhancement in GB levels was more

pronounced at anthesis (104.8 %) as compared to maxi-

mum tillering stage (77.0 %). Application of GB to

drought stressed plots significantly increased its own con-

tent. Further, this response of GB sprayed plots was con-

sistent across the genotypes at both the maximum tillering

(22.3 %) and anthesis stages (24.4 %). Interestingly, higher

levels of GB in sprayed plots at anthesis were in contrast to

reduction observed in choline accumulation at that stage.

Plant Growth Regul (2014) 72:221–228 223

123

Sucrose content

Drought stress caused a significant increase in sucrose

content of stressed plots over irrigated ones (Fig. 4a, b).

This enhancement in levels of sucrose under drought was

greater at anthesis (46.4 %) than at maximum tillering

(39.6 %) stage. Application of GB significantly decreased

the sucrose content. Moreover, the magnitude of reduction

in GB sprayed plots was higher at anthesis (-42.0 %) than

at maximum tillering stage (-18.3 %). This response in

sucrose levels of sprayed plots was consistent across the

genotypes.

0

1

2

3

4

5

6

7

8

9

10Pr

olin

e co

nten

t (µm

oles

/gFW

)

Genotype

ControlStressStress+GB

(a)

0

1

2

3

4

5

6

7

8

9

10

Prol

ine

cont

ent (

µmol

es/g

FW)

Genotype

Control

Stress

Stress+GB

(b)

Fig. 1 Proline content of wheat genotypes as affected under drought

stress and with GB application in drought stress conditions. a Max-

imum tillering stage: G = 0.605, T = 0.240 G 9 T = 1.048.

b Anthesis stage: G = 0.348, T = 0.138, G 9 T = 0.602 where G

and T corresponded to genotypes and treatments and G 9 T were

genotypes and treatments interactions

0

2

4

6

8

10

12

14

16

18

20

Cho

line

cont

ent (

µmol

es/g

DW

)

Genotype

Control

Stress

Stress+GB

(a)

0

2

4

6

8

10

12

14

16

18

20

Cho

line

cont

ent (

µmol

es/g

DW

)

Genotype

ControlStressStress+GB(b)

Fig. 2 Choline content of wheat genotypes as affected under drought


imum tillering stage: G = 0.381, T = 0.151, G 9 T = 0.659.




0

2

4

6

8

10

12

GB

con

tent

(µm

oles

/gD

W)

Genotype


(a)

0

2

4

6

8

10

12

GB

con

tent

(µm

oles

/gD

W)

Genotype

Control

Stress

Stress+GB(b)

Fig. 3 Glycine betaine levels in leaf tissues as affected under drought


imum tillering stage: G = 0.735, T = 0.292, G 9 T = 1.273.





123

Relative leaf water content and stress tolerance index

As represented in Table 1, relative leaf water content

(RWC) was significantly higher in irrigated (control) plots

over their drought stressed counterparts. GB application

was found to improve RWC of wheat genotypes under

drought stress at both the stages but short of the level

observed in irrigated plots. Moreover, STI of sprayed plots

was also found to be significantly increased over non-

sprayed counterparts (Table 1). In order to evaluate the

association of RWC and STI to levels of compatible solutes

under drought stress, a correlation analysis was carried out

0

5

10

15

20

25

30

35

40

45Su

cros

e co

nten

t (m

g/gF

W)

Genotype


(a)

0

5

10

15

20

25

30

35

40

45

Sucr

ose

cont

ent (

mg/

g FW

)

Genotype


(b)

Fig. 4 Sucrose accumulation in leaf tissues under drought stress and

with GB application in drought stress conditions. a Maximum

tillering stage: G = 4.029, T = 1.601, G 9 T = 6.978. b Anthesis

stage: G = 2.039, T = 0.810, G 9 T = 3.532 where G and T

corresponded to genotypes and treatments and G 9 T were genotypes

and treatments interactions

Table 1 Comparison of relative water content and stress tolerance index of wheat genotypes subjected to irrigation (control), drought stress and

exogenous application of glycine betaine (100 mM) under drought stress conditions

Genotypes Relative water content (%) Stress tolerance index

Maximum tillering stage Anthesis stage

Irrigated Drought

stress

Drought

stress ? GB

Irrigated Drought

stress

Drought

stress ? GB

Drought

stress

Drought

stress ? GB

C306 86.071 77.778 80.619 82.846 78.000 80.118 0.539 0.782

PBW175 85.769 74.648 82.615 84.375 78.392 80.000 0.572 0.736

PBW527 83.063 71.233 80.000 81.481 78.356 81.673 0.604 0.790

PBW343 87.732 80.000 83.117 84.127 80.636 82.792 0.455 0.673

PBW550 85.964 83.019 82.483 82.500 79.310 80.982 0.809 0.820

DBW17 83.651 80.060 80.615 82.049 76.667 78.500 0.627 0.850

BW9022 83.750 81.441 81.571 81.136 79.697 80.484 0.415 0.687

BW9097 84.118 79.688 80.000 84.211 78.275 80.000 0.495 0.763

BW9183 84.030 81.885 80.667 81.967 78.185 80.714 0.676 0.922

BW9005 86.091 81.500 82.690 86.228 79.655 80.240 0.995 1.078

BW9016 85.455 80.625 82.500 87.710 80.556 81.143 0.526 0.918

BW8989 86.189 82.060 83.333 85.075 80.848 82.235 0.924 1.013

BW8366 87.324 71.642 84.714 87.831 78.207 80.462 0.935 0.913

BW9151 83.846 73.333 81.507 82.645 79.929 81.037 0.634 0.680

BW8362 86.302 70.423 82.545 84.906 77.186 78.846 0.703 0.931

BW 9025 86.873 81.159 82.235 83.134 77.090 79.323 0.572 0.621

BWL0089 87.500 83.333 84.103 87.356 78.484 80.714 0.655 0.791

PBW621 86.041 75.000 80.011 85.000 78.846 82.791 0.943 1.095

HD2967 83.151 80.211 82.014 82.258 78.214 81.667 0.926 1.031

* CD at 5 % level of significance for RWC: Maximum tillering stage: G = 3.626, T = 1.441, G 9 T = 6.281 For STI: G = 0.154, T = 0.501,

G 9 T = NS

Anthesis stage: G = NS, T = 1.679, G 9 T = NS; where CD represented critical differences at 5 % level of significance, NS corresponded to

non significant differences, G corresponded to genotypes, T corresponded to treatments and G 9 T corresponded to interaction between

genotypes and treatments


123

under both non-sprayed and GB sprayed conditions

(Table 2). Increased levels of proline, choline and sucrose

content as observed in drought stressed plots at maximum

tillering stage were not correlated significantly to levels of

RWC under stress conditions. Further this non-significant

relationship of above mentioned osmolytes was also

observed at anthesis stage. Following a similar correlation

pattern, contribution of osmolytes (proline, choline and

sucrose) to STI was non-significant at both the stages under

drought stress.

As mentioned previously, GB application induced

changes in levels of above mentioned osmolytes which

showed stage-specific and genotype dependent behaviour.

Inspite of the changes, their correlation with RWC and STI

was observed to be non-significant in GB sprayed plots.

Interestingly, GB content at maximum tillering stage

showed significant correlation with RWC and STI to a

greater extent in comparison to other osmolytes. GB con-

tent of drought stressed plots at maximum tillering stage

showed significant negative correlation with STI (r =

-0.533). Reversal of this relationship occurred with GB

spray at maximum tillering stage. This negative correlation

between GB and STI, at maximum tillering suggested that

enhanced levels of GB at early stages under drought stress

could decrease the tolerance ability. No such response

between GB content and STI was observed at anthesis

under drought stress. An enhanced level of GB in sprayed

plots at the stage of maximum tillering was also signifi-

cantly negative correlated with RWC (r = -0.580). In

contrast, GB levels of non sprayed plots at maximum til-

lering stage and GB levels at anthesis under both sprayed

and non sprayed conditions were not correlated with RWC.

While correlation was studied between RWC and STI at

both stages, non significant relationship existed between

the two at both stages.

Discussion

Osmolyte accumulation is often, related with osmotic

adjustment, without knowing the contribution of osmolytes

to this physiological mechanism. Moreover, maintenance

of turgor and dehydration tolerance has also been associ-

ated with the phenomenon of OA. All these strategies were

thought to enhance plant resistance to water deficit as well

as other abiotic stresses. Based on this, the present research

was carried out to study the variations in content of various

osmolytes in wheat genotypes under drought stress and

their relationship with drought tolerance in wheat. Fur-

thermore, exogenous application of GB was done at

selected growth stages i.e. maximum tillering and anthesis,

to study the effect of this osmolyte on its own content and

contents of other compatible solutes produced under

drought stress conditions. This osmolyte was selected on

basis of previous studies on exogenous application of

osmolytes, which supported enhancement in tolerance of

wheat crop to different abiotic stresses via osmotic

adjustment, improved anti-oxidative defense system and

increased photosynthetic rate by GB (Ma et al. 2006; Raza

et al. 2007) which could result in yield benefits.

In current study, foliar application of GB (100 mM)

significantly improved STI of wheat genotypes under

drought stress. In order to determine whether this

improvement was because of OA in GB treated plots,

contents of various osmolytes were estimated. GB appli-

cation under drought stress conditions resulted in enhanced

levels of osmolytes (proline and GB) but lowering the

sucrose content in leaves. This decrease in sucrose content

could be due to involvement of sucrose in GB induced

synthesis of compatible solutes like proline, choline etc., by

either acting as a carbon skeleton or as an energy source.

Another reason would be more transport of sucrose to

promote growth of developing sinks such as roots at til-

lering and grains at anthesis under drought stress by GB. In

contrast to enhanced levels of proline and GB due to GB

application, choline showed a decline at anthesis stage.

This was observed across all 19 wheat genotypes. Choline

Table 2 Correlation analysis of relative water content (RWC) and

stress tolerance index (STI) with various osmolytes under drought

stress and exogenously applied glycine betaine (GB) conditions

Parameters RWC STI

Maximum

tillering

Anthesis Maximum

tillering

Anthesis

Drought stress

Proline -0.320 -0.254 0.160 -0.169

Choline 0.404 0.116 -0.332 0.140

Glycine

betaine

0.346 -0.338 -0.533* -0.004

Sucrose 0.408 -0.275 -0.064 -0.010

STI -0.063 0.035 1.000 1.000

Drought stress ? GB

Proline 0.079 0.071 0.137 -0.225

Choline -0.240 -0.026 -0.164 0.233

Glycine

betaine

-0.580* 0.368 0.001 0.218

Sucrose -0.077 0.110 0.074 0.262

STI 0.035 0.243 1.000 1.000

* Significant at 5 % level of significance. Significant correlation

(P B 0.05) suggested the dependence of one component on the other

or in other terms as one component determined the level of other

component. Moreover, significant negative correlation indicated that

the studied components were related to each other in a way that

increase in one component decreases the value of other component.


123

is a precursor of GB and reduction in its levels due to

exogenous GB application may be explained by presence

of a feed-back loop. Significant increase in RWC and STI

was observed in case of GB sprayed plots which seemed

due to OA in these wheat genotypes. But non-significant

correlation of osmolytes levels (such as proline, choline

and sucrose) with RWC and STI, at both the stages under

GB sprayed and non sprayed drought stressed conditions

indicated that production of these osmolytes could be a

consequence of stress rather than a metabolic response.

Moreover GB content of sprayed plots and RWC were

negatively correlated at maximum tillering stage. No

relationship between them was observed at anthesis.

Interestingly, GB content of drought stressed plots at

maximum tillering stage was negatively correlated with

STI which suggested that enhancement in GB levels could

decrease the tolerance index, thus may cause yield reduc-

tions under stress. This was in contrast to results obtained

with GB spray under drought stress where enhanced levels

of GB and increased STI were observed, although no

correlation existed between them. The reason behind this

response of GB to STI under non-sprayed and GB sprayed

condition is unclear. It could be hypothesized that accu-

mulated osmolytes might be involved in some downstream

metabolic processes of signal transduction under drought

stress. There may be a possibility that GB accumulation

could have some indirect effects on STI via changes in

sucrose metabolism etc., under drought stress which in turn

affected dry matter accumulation. Previous studies on

maize (Bolanos and Edmeades 1991) and barley (Grumet

et al. 1987) also reported that correlation between OA and

performances under drought were weak, inconsistent and

non-significant. Our results supported reviewed studies on

OA by Serraj and Sinclair (2002), who suggested that

under natural stress environments, stress is not lethal to

affect plant survival where osmoprotection could play a

role. OA could cause osmoprotection, turgor maintenance

and dehydration avoidance at severe stress level which

might threaten plant survival. Such severe stress levels

could result in loss of crop yield capability and crop yields

will be low. If OA operate in leaves, resulting in turgor

maintenance causing leaf area expansion, delay in leaf

wilting and rolling, delayed stomatal conductance, then

there would be increased risk of exhausting soil water

under drought stress and high rates of plant dehydration

would be sustained. This would result in leaf and plant

death when the threshold level of lethal RWC reaches.

Thus in conclusion, our results suggested that OA in

leaves was not responsible in causing tolerance to drought

and for yield benefits in wheat under drought stress. So OA

could not be considered as a selection criteria for drought

tolerance in wheat. There is a need for a more precise

research on osmotic adjustment via these compatible

solutes and their exogenous application to alleviate various

abiotic stresses.

Acknowledgments This work was supported by Council of Scien-

tific and Industrial Research (Project No. 09/272/0129-EMR-I), New

Delhi, India.

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Glycine betaine application modifies biochemical attributes of osmotic adjustment in drought stressed wheat