Annals of Botany 102: 287 –293, 2008
doi:10.1093/aob/mcn091, available online at www.aob.oxfordjournals.org
The Effect of Temperature Shock and Grain Morphology on Alpha-amylase
in Developing Wheat Grain
A . D . FAR R E L L and P. S . K E T T L E W E L L*
Crop and Environment Research Centre, Harper Adams University College, Newport, Shropshire, TF10 8NB, UK
Received: 14 December 2007 Returned for revision: 8 April 2008 Accepted: 12 May 2008 Published electronically: 4 June 2008
† Background and Aims The premature production of alpha-amylase without visible germination has been observed
in developing grain of many cereals. The phenomenon is associated with cool temperatures in the late stages of grain
growth but the mechanisms behind it are largely unknown. The aim of this study was to replicate the phenomenon
under controlled conditions and investigate the possibility of a mechanistic link with grain size or endosperm cavity
size.
† Methods Five wheat (Triticum aestivum) genotypes differing in their susceptibility to premature alpha-amylase
were subjected to a range of temperature shocks in controlled environments. A comparison was then made with
plants grown under ambient conditions but with grain size altered by using degraining to increase the assimilate
supply. At maturity, alpha-amylase, grain area and endosperm cavity area were measured in individual grains.
† Key Results Both cold and heat shocks were successful in inducing premature alpha-amylase in susceptible genotypes, with cold shocks the most effective. Cold shocks also increased grain area. Degraining resulted in increased
grain area overall, but the larger grain did not have higher alpha-amylase. Analysis of individual grain found that
instances of high alpha-amylase were not associated with differences in grain area or endosperm cavity area.
† Conclusions Pre-maturity alpha-amylase is associated with temperature shocks during grain filling. In some cases
this coincides with an increase in grain area, but there is no evidence of a mechanistic link between high alphaamylase and grain or endosperm cavity area.
Key words: Alpha-amylase, pre-maturity alpha-amylase, late maturity alpha amylase, temperature, grain size, endosperm
cavity, wheat, Triticum aestivum.
IN TROD UCT IO N
The role of alpha-amylase in germination is well studied;
less well characterized is the phenomenon of premature
alpha-amylase. Pre-maturity alpha-amylase (PMA, also
called late maturity amylase) refers to the sporadic production of alpha-amylase during the later stages of grain
filling. This genetic defect is frequently detected in commercially grown wheat and other cereals (Lunn et al.,
2001). Although it was initially associated with pre-harvest
sprouting, PMA is not necessarily accompanied by sprouting and is now thought to be under independent genetic
control (Mares and Mrva, 2008).
In some genotypes PMA appears to be constitutively
expressed while in others the occurrence is sporadic
(Mares and Mrva, 2008). In the field, PMA is often associated with cool, wet conditions during grain development
(Gale et al., 1987; Mrva and Mares, 1994). PMA has also
been detected in controlled environment experiments
when temperature shocks are applied during grain filling
and it has been proposed that there is a subset of genotypes
that require a temperature shock to express PMA (Mares
and Mrva, 2008). Cold shocks have been effective in inducing PMA in Australian genotypes, but similar experiments
with UK genotypes have proved less reliable (Gale et al.,
1987; Major, 1999; Tjin-Wong-Joe, 2004). Heat shocks
have also been found to stimulate PMA in some cases
* For correspondence. E-mail [email protected]
(Randall and Moss, 1990; Major, 1999). The variability
in results is partially explained by the observation of
Mares and Mrva (2008) that a temperature shock is only
effective if applied during a ‘window of sensitivity’,
which extends from 25– 30 d after anthesis under typical
Australian conditions.
One limitation in relating such results to field-grown
plants is the lack of a mechanistic model linking variation
in temperature to alpha-amylase production. A central question is whether temperature shocks act directly on
alpha-amylase-producing cells or indirectly through their
effect on grain morphology. Although both temperature
and grain morphology have been proposed as PMAregulating factors the link between the two has not been
investigated.
Evers et al. (1995) and Evers (2000) reviewed the evidence for a link between large grain size and high
alpha-amylase activity. The evidence presented follows
two lines: firstly, of the varieties commonly grown in the
UK those classed as large-grained were all in the high
alpha-amylase (low Hagberg falling number) category; secondly, some studies have found a gradient in alpha-amylase
activity within ears and within spikelets that correlates
with the typical weight distribution of individual grains.
Although not conclusive, these studies suggest a fundamental mechanistic link between grain size and alpha-amylase
that is expressed both within and between genotypes. The
studies also suggest that it is external grain dimensions
(grain area) rather than grain weight that is most associated
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Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
with PMA. Kindred et al. (2005) also found a correlation
between grain area and alpha-amylase. They suggested
that the underlying relationship is between alpha-amylase
and the size of the endosperm cavity, with grain area
acting as an imperfect proxy. As also seen by Evers
(2000), the correlation occurred both between genotypes
and within genotypes grown at different nitrogen levels.
The proposal that PMA might be associated with the size
and disruption of the endosperm cavity relates to observations that alpha-amylase produced during PMA tends to
be localized around the grain crease, possibly being produced in the aleurone layer surrounding the endosperm
cavity (Cornford et al., 1987; Evers et al., 1995;
Greenwell et al., 2001; Tjin-Wong-Joe, 2004; Kindred
et al., 2005). Again, there is some disagreement between
UK studies and those carried out in Australia where PMA
was found to have a random distribution throughout the
aleurone (Mrva et al., 2006; Mares and Mrva, 2008). The
relationship between cavity size and temperature shocks
has not been investigated.
Here, we apply a range of temperature shocks and look at
the effect on alpha-amylase and grain morphology in
several UK winter wheat genotypes. A comparison is
made with plants grown under ambient temperatures but
with grain morphology altered by using degraining to
increase the assimilate supply.
M AT E R I A L S A N D M E T H O D S
Plant material
Seeds of the UK winter wheat (Triticum aestivum L.) varieties ‘Maris Huntsman’, ‘Rialto’, ‘Spark’, ‘Option’ and
‘Potent’ were obtained from J. Flintham (John Innes
Centre, Norwich, UK). Based on results from the field,
‘Maris Huntsman’, ‘Rialto’ and ‘Spark’ represent high,
medium and low susceptibility to PMA respectively
(J. Flintham, pers. comm.), while the remaining genotypes
are less well characterized. ‘Maris Huntsman’ and ‘Spark’
were chosen to provide within-treatment controls for the
complex temperature regimes where it was not feasible to
include other control treatments.
TA B L E 1. Outline of day/night temperatures (8C) and timing
of treatments in the temperature-shock experiments
Treatment stage
Treatment
Pre-anthesis
Pre-treatment
Induction*
Ripening
Mid ! Low
Mid ! High
High ! Mid
Glasshouse
Glasshouse†
Glasshouse
22/22
20/10
30/15
12/12
30/20
18/12
22/22
30/15
30/15
*Induction started at 550 degree-days after anthesis and lasted 8
calendar days.
†
Mid ! High planted in a glasshouse on 25 May 2006 and transferred
during June 2006. Other treatments were planted in a glasshouse on 2
Dec. 2005 and transferred during Feb. 2006.
practical reasons experiments were carried out in two
stages at different times of year (see Table 1 for details).
Plants were checked regularly and tagged at anthesis.
When a sufficient number of plants reached anthesis on
the same day, this subset of plants was transferred to one
of three cabinets. This ensured that all plants used were at
a comparable developmental stage. Each cabinet temperature was initially set to be similar to that of the glasshouse.
Over the next 12 d, temperatures were gradually adjusted
until a defined pre-treatment temperature was reached.
Three cabinets were used to apply three temperature
shocks: Mid ! Low, Mid ! High, and High ! Mid, as
shown in Table 1. The third temperature shock, ‘High !
Mid’, was designed to replicate previous experiments
carried out under Australian conditions (Mrva and Mares,
2001). In each case the temperature shock started approximately 550 degree-days after anthesis (degree-DAA, base
temperature 0 8C) and lasted for 8 d. The start time
ranged from 25 to 30 DAA depending on the pre-treatment
regime. Plants remained in the cabinets until 60 DAA when
the grain was harvested. The cabinets were maintained on a
14/10 h light/dark cycle with a relative humidity of 70 %,
photosynthetic photon flux density was 250– 300 mmol
m22 s21 throughout and pots were kept near field capacity
using an automatic drip-irrigation system attached to
each pot.
Degraining
Temperature shock
Plants were vernalized at 6 8C for 8 weeks then transferred to 1.3-L pots in a glasshouse ( pots contained John
Innes No. 2 Compost: Keith Singleton’s Seaview
Nurseries, Cumbria, UK). Secondary shoots (tillers) were
removed periodically to leave the main shoot plus four secondary shoots. Plants were reared in the glasshouse until
anthesis then transferred to controlled-environment cabinets
(Fitotron, Sanyo Gallenkamp, Loughborough, UK; or
Conviron, Controlled Environments, Winnipeg, Canada).
The glasshouse was heated to provide a minimum of
15/5 8C day/night temperature, water was provided by
capillary matting wetted automatically three times a day,
and supplementary light (400 W high pressure sodium)
was used to maintain a minimum 16-h day length. For
Plants were grown from seed in a polytunnel where they
underwent vernalization at ambient temperatures over
winter. Secondary shoots were removed periodically to
leave the main shoot plus four secondary shoots.
At anthesis half of the plants were left intact while half
were subjected to 50 % spikelet removal (degrained) in
order to stimulate compensatory grain growth in the remaining grains. Degraining was achieved by excising an entire
row of spikelets from one side of the main ear and removing
the whole ear from all secondary shoots.
For three genotypes, ‘Maris Huntsman’, ‘Spark’ and
‘Rialto’, the degraining treatment was combined with a
cold shock applied in controlled-environment cabinets (as
above). Plants were transferred to the cabinets at anthesis
and underwent either an ‘ambient’ treatment (similar to
Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
TA B L E 2. Outline of day/night temperatures (8C) and timing
of treatments in the degraining experiments
Treatment stage
Treatment
Pre-anthesis
Pre-treatment
Induction*
Ripening
Ambient†
Mid ! Low (B)
Polytunnel
Polytunnel
25/15
25/15
25/15
13/11
Polytunnel
Polytunnel
*Induction started at 330 degree-days after anthesis and lasted 15
calendar days.
†
‘Option’ and ‘Potent’ remained in the polytunnel from planting
(26 Dec. 2005) until maturity and are also classed as ambient.
the temperatures in the polytunnel) or a Mid ! Low (B)
treatment, as described in Table 2. At 28 DAA both
control and Mid ! Low (B) plants were returned to the
polytunnel. The cabinets were maintained on a 16/8 h
light/dark cycle with a humidity of 70 % and a photosynthetic photon flux density of 500 mmol m22 s21 throughout.
Grain alpha-amylase assay
All grain was sampled at random from within a target
region consisting of florets 1 and 2 from spikelets 7 to 11
(counting acropetally) of the main shoot. For each plant
one grain was removed, weighed and an image taken of
the grain area ( plan view of ventral side) and transverse
section (following hand-dissection with a scalpel). Images
were acquired with a flat-bed scanner (ScanExpress,
Mustek, Taiwan) and analysed using ImagePro Plus
(Media Cybernetics, Marlow, UK). Transverse sections
were used to measure the area of the endosperm cavity
(Fig. 1).
1 mm
289
High pI alpha-amylase protein content was assayed
using a high pI-specific ELISA (Value Added Wheat
CRC, North Ryde, NSW, Australia; Mrva and Mares,
2001). The alpha-amylase assay was carried out using a
single half-grain from each plant. The distal half of each
grain was reduced to 20 + 2 mg before being homogenized
for use in the assay. Grain halves were homogenized in
microcentrifuge tubes using a rotating ball grinder
(RotoMix, 3M ESPE AG, Seefeld, Germany) and 0.5 mL
of extraction buffer (85 mmol NaCl) was added to each
tube. Tubes were incubated for 1 h at room temperature
on a shaker, after which 100 mL of grain extract was
taken for use in the assay. Absorbance was measured at
450 nm using a microplate reader (Bio-Rad, Benchmark,
Watford, UK) and results zeroed against extraction
buffer blanks.
Statistical analysis
The data presented are means of eight grains per treatment with each grain taken from one of eight replicate
plants. Statistical analysis was carried out using two-way
ANOVA with multiple comparisons (l.s.d., P , 0.05)
using Genstat 8 (VSN International Ltd., Hemel
Hempstead, UK). For alpha-amylase, log-transformed data
was also tested but was found to show similar output to
untransformed data.
R E S U LT S
Temperature shock
Environment, genotype and environment genotype all
produced significant differences in alpha-amylase. Grain
area showed a similar response, although the
environment genotype effect was not significant.
‘Spark’, a PMA-resistant genotype, produced negligible
levels of alpha-amylase in all treatments (Fig. 2). The
Mid ! Low and the Mid ! High temperature shock
stimulated alpha-amylase production in ‘Maris Huntsman’
and ‘Rialto’. The High ! Mid temperature shock, which
was designed to replicate previous experiments carried
out under Australian conditions (Mrva and Mares, 2001),
resulted in negligible alpha-amylase levels in all three
genotypes.
Across the three treatments there was a clear correlation
between alpha-amylase content and grain area, with
cooler temperatures over the grain-filling period resulting
in larger grains with more alpha-amylase (Fig. 2). A
similar correlation was also present within treatments.
Degraining
43 mg
64 mg
F I G . 1. Typical grain images for ‘Rialto’, showing the range in grain
weight, grain area and endosperm cavity area.
In the degraining experiment, Mid ! Low (B) successfully stimulated alpha-amylase in intact ‘Rialto’ plants
(Fig. 3). ‘Maris Huntsman’ produced high alpha-amylase
levels in both ambient and Mid ! Low (B) treatments.
As in Fig. 2, a positive relationship was found between
grain area and alpha-amylase for ‘Maris Huntsman’,
290
Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
Alpha-amylase (OD)
0·50
‘Spark’
‘Rialto’
‘Huntsman’
A
0·45
0·40
0·35
0·30
0·25
0·20
0·15
0·10
0·05
0·00
26
B
Grain area (mm2)
24
22
Grain and cavity size
Figure 4 shows grain area, cavity area and alpha-amylase
content for individual grains, with data combined from all
of the treatments used in the degraining experiments.
Alpha-amylase appears to show a weak positive correlation
with grain area when genotypes are ignored. However, a
regression analysis with genotypes as groups shows that
the association is due to differences between genotypes.
Within genotypes there is no association between
alpha-amylase and grain or cavity area.
Any cavity with an area more than 1.5 s.d. from the mean
(i.e. .0.6 mm2) is classed as a ‘large cavity’. ‘Maris
Huntsman’ and ‘Option’ account for the majority of
instances of large cavities; these instances were not associated with the degraining or Mid ! Low (B) treatments
(data not shown). Neither is there a higher incidence of
high alpha-amylase content among those grains with
‘large cavities’ (Fig. 4A).
20
DISCUSSION
18
16
HighÆMid
MidÆHigh
MidÆLow
F I G . 2. (A) Grain alpha-amylase content and (B) grain area of mature
winter wheat subjected to different temperature regimes during grain
filling (temperature-shock experiments; see details in Table 1). Data are
means of eight replicate plants. Vertical bars indicate l.s.d.005 for
genotype environment.
‘Rialto’ and ‘Spark’; however, this correlation was not seen
in the two additional genotypes ‘Option’ and ‘Potent’
(Fig. 2).
In the intact plants, environment, genotype and
environment genotype all produced significant differences in alpha-amylase. Grain area showed a similar
response, although the environment genotype effect was
not significant. In the degrained plants, only genotype had
a significant effect on alpha-amylase. Grain area again
showed significant differences due to environment and
genotype but not environment genotype. Degraining
resulted in significantly larger grains (Fig. 3), when averaged over all five genotypes grown under ambient conditions. The increased grain area did not, however,
produce significant changes in alpha-amylase content.
Combining degraining and Mid ! Low (B) treatments
had an additive effect on grain area, producing larger
grains than either treatment on its own (Fig. 3). Again,
the increased grain area was not associated with an increase
in alpha-amylase. In fact, the expected increase in
alpha-amylase seen in ‘Rialto’ following the Mid ! Low
(B) treatment was absent in the degrained plants (Fig. 3).
The Mid ! Low (B) treatment did stimulate PMA in
intact ‘Rialto’ plants, but the lower levels of alpha-amylase
overall suggest this treatment was less effective than the
previous Mid ! Low treatment that was applied at a later
stage of grain filling.
The understanding of pre-maturity alpha-amylase (PMA)
has often been hindered by the sporadic way in which it
occurs, with levels of alpha-amylases varying widely
between plants, between ears and between grain. The
experiments here rely on a relatively small sample size
(eight grain, each from a replicate plant), with grain taken
from a target region thought to have the highest occurrence
of PMA (Gale et al., 1987). Adopting this method and analysing the results on a single-grain basis appears to provide
a good system for probing the mechanisms behind PMA.
Temperature shock
These results confirm that ‘Maris Huntsman’ is highly
susceptible to PMA while ‘Spark’ appears resistant.
‘Rialto’ falls into an intermediate category, with PMA
only expressed under particular growth conditions. As
such, ‘Rialto’ provides a suitable model to test the possible
link between alpha-amylase and grain or endosperm cavity
area. It is this intermediate category that is of most interest
here, as it gives insight into the physiological processes
underlying PMA.
Of the three temperature regimes applied here, Mid !
Low was the most effective in inducing PMA. The Mid
! Low (B) treatment applied in the degraining experiments (Fig. 3) confirmed the effectiveness of a cold
shock in stimulating the production of alpha-amylase.
This is consistent with results of similar cold-shock experiments using Australian wheat genotypes (Mares and Mrva,
2008). The Mid ! High regime was also effective in inducing PMA, suggesting that a heat shock has a similar effect.
The negligible levels of alpha-amylase in all three genotypes after the third temperature shock ‘High ! Mid’ was
surprising given that this represented a similar temperature
change to the other treatments and was applied at a similar
developmental stage (550 degree-DAA). Nevertheless
this attempt to replicate Australian conditions involved
applying a wide range of temperatures pre-anthesis and
Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
Degrained ears
Intact ears
0·50
A
B
C
D
‘Spark’
‘Rialto’
‘Huntsman’
‘Option’
‘Potent’
0·45
0·40
Alpha-amylase (OD)
291
0·35
0·30
0·25
0·20
0·15
0·10
0·05
0·00
26
Grain area (mm2)
24
22
20
18
16
Ambient
Mid Æ Low (B)
Ambient
Mid Æ Low (B)
F I G . 3. (A, B) Grain alpha-amylase content and (C, D) grain area of intact and degrained winter wheat subjected to different temperature regimes during
grain filling (degraining experiments; see details in Table 2). Data are means of eight replicate plants. Vertical bars indicate l.s.d.005 for genotype environment.
0·50
A
B
0·45
Alpha-amylase (OD)
0·40
0·35
0·30
0·25
0·20
‘Spark’
‘Rialto’
‘Huntsman’
‘Option’
‘Potent’
0·15
0·10
0·05
0·00
0·0
0·2
0·4
0·6
0·8
1·0
2
Cavity area (mm )
1·2
1·4
1·6
14·0
16·0
18·0 20·0 22·0 24·0
Grain area (mm2)
26·0
28·0
30·0
F I G . 4. The relationship between alpha-amylase content and (A) endosperm cavity area and (B) grain area. Grain to the right of the line in (A) are
designated as having a ‘large cavity’. Data are for single grains grown under a range of conditions as indicated in Fig. 3.
292
Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
pre-treatment, which can reduce the effectiveness of the
cool shock (Mares and Mrva, 2008).
Temperature influences many aspects of grain development, not least the rate and duration of grain filling. In
general, wheat has an optimum temperature for grain
filling of 15 8C, with each 1 8C rise above this resulting
in a 3 – 5 % reduction in single grain weight (Wardlaw
et al., 1989). In the temperature-shock experiment the
temperatures varied over the grain-filling period, but
overall the average temperature applied in each regime
(Table 1) showed a negative association with grain area
(Fig. 2). The tendency for cool temperatures during grain
filling to produce both larger grain and increase PMA
may explain the association found between the two traits
in field-grown plants.
As lower temperatures increase both grain area and PMA
the possibility of a mechanistic link between the two
emerges. It is possible that cold shocks and cool periods
in the field induce PMA by altering grain filling, e.g. by
decoupling the growth of the aleurone layer from the expansion of the endosperm or endosperm cavity. This possibility
is supported by the positive association found between
grain size and PMA in other circumstances (Evers, 2000;
Kindred et al., 2005). Taken in isolation the results of the
temperature-shock experiment are consistent with this proposition, although the results from the degraining experiment
appear to counter this. In the degraining experiment the
association between grain area and PMA was lost. Given
this, and the fact that both cold and heat shocks can be
effective, it is more likely that temperature affects PMA
through other means.
Grain area and PMA
The degraining experiment was designed to test if there
was a causal link between grain area and PMA as suggested
by other studies (Evers et al., 1995; Evers, 2000; Greenwell
et al., 2001). Degraining generally increases grain size by
reducing the number of grains competing for photoassimilates and stimulating compensatory grain growth in the
remaining grain (Borras et al., 2004). For the four genotypes that showed a positive response to degraining there
was an average increase in grain area of 11 %. This is
lower than has been seen in some studies but comparable
to the 11 % expected for wheat in general (Borras et al.,
2004). Despite the increase in grain area there was no
general increase in alpha-amylase. Only ‘Potent’ showed
an increase in both traits with the alpha-amylase level
remaining relatively low throughout. Similarly, the
between-genotype association of grain area and PMA
seen in the temperature-shock experiment and by Evers
(2000) was not seen when all five genotypes were tested.
Similar results were seen when grain weight rather than
area was used (data not shown). Even though this comparison of five genotypes is far from comprehensive, the fact
that ‘Potent’ and ‘Option’ have grain areas equal to or
greater than ‘Maris Huntsman’, yet have consistently
lower levels of alpha-amylase, indicates that grain area is
not a reliable predictor of PMA. This is confirmed by the
analysis of individual grains, with many instances of
PMA detected in grain of average area (Fig. 4B).
Although it remains possible that particular cases of
enlarged grain or particular genetic determinants of grain
size also influence PMA, our results do not support
the suggestion of a mechanistic relationship between the
two traits.
Endosperm cavity area and PMA
Kindred et al. (2005) found that reducing the amount of
applied N resulted in increased alpha-amylase and endosperm cavity size across several genotypes. In addition
to this indirect association, when comparing ‘large-cavity
grains’ to ‘small-cavity grains’ they found a similar positive
relationship between cavity size and alpha-amylase within
genotypes grown under the same N level. In the present
work, there was a weak association between grain area
and endosperm cavity area (data not shown) and grain
area showed a weak association with alpha-amylase
content (Fig. 4). However, despite the range of
alpha-amylase levels detected, no clear relationship with
cavity area was seen under any of the treatments (Fig. 4).
A similar lack of association was seen whether cavity size
was expressed as area or maximum diameter (data not
shown). The different results may reflect differences in
the methods used to measure alpha-amylase; Kindred
et al. (2005) measured alpha-amylase levels by placing
half-grains transverse-side down on agar and measuring
the extent of starch degradation. It may be that differences
in cavity size alter the amount of alpha-amylase reaching
the agar without altering the overall alpha-amylase level
in the homogenized grain (as measured here).
Thus, neither total grain area nor endosperm cavity area
explained the variation in PMA seen here. Observations
of PMA occurrence in tall and semi-dwarf genotypes
have also revealed no obvious link with grain morphology,
indicating that significant changes in the GA signal transduction pathway are not involved. Nonetheless, production
of alpha-amylase in the aleurone is GA dependent and
GA-insensitive genotypes are generally less prone to
PMA than their tall parents (Mares and Mrva, 2008). It is
likely that GA and GA-sensitivity are key factors in stimulating PMA here. The most effective treatment (Mid !
Low) was applied in the final stages of grain filling as the
grain was approaching physiological maturity. In vitro
experiments have shown that in the presence of GA,
aleurone tissue from such immature grain can be induced
to synthesise alpha-amylase when subjected to a suitable
stimulus. Successful stimuli have included low temperature,
high temperature, drying and incubation in a buffer (Bewley
and Black, 1994). Given the range of treatments that can
induce alpha-amylase in detached grain, it has been
suggested that a regulatory mechanism exists in vivo to
inactivate the GA signal, or make the aleurone layer insensitive to GA (Cornford and Black, 1985). One candidate for
this regulatory mechanism is the GA/ABA ratio. Peak ABA
levels also occur in the later stages of grain filling as maturation begins (King, 1976; Finkelstein, 2004). Applying a
temperature shock during the window of sensitivity could
stimulate PMA by disrupting this regulatory mechanism,
Farrell and Kettlewell — Temperature, Grain Morphology and Alpha-amylase in Wheat
e.g. by altering the GA/ABA ratio itself or the sensitivity of
the aleurone to the two hormones.
AC KN OW L E DG E M E N T S
We thank Tony Evers for instructive comments on the manuscript, Daryl Mares and Kolumbina Mrva for assistance in
developing methods, Janice Haycox for technical assistance,
and James Simmonds and John Flintham for providing plant
material. This work was sponsored by the Department
of Environment, Food and Rural Affairs, and the
Biotechnology and Biological Sciences Research Council
through the Sustainable Arable LINK Programme, and supported by Home Grown Cereals Authority.
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