Journal of Experimental Botany, Vol. 49, No. 326, pp. 1519–1528, September 1998
Effects of alien cytoplasmic variation on carbon
assimilation and productivity in wheat
Peter Jones1,3, Eavan M. Keane1 and Bruce A. Osborne2
1Department of Plant Science, University College Cork, Cork, Ireland
2Department of Botany, University College Dublin, Belfield, Dublin 4, Ireland
Received 2 January 1998; Accepted 30 April 1998
Abstract
In glasshouse studies of four alloplasmic wheat series,
phenotypic characters were least affected when the
recipient parent cytoplasm was replaced by donor
cytoplasm of the S or D plasmatype. In the T. aestivum
cv. ‘Selkirk’ series, cytoplasm substitution did not
affect P
per unit leaf area, although the flag leaf
max
area (and photosynthetic rate per leaf ) of each alloplasmic line was greater than that of euplasmic
‘Selkirk’. In field trials, all the D plasmatype alloplasmics tested produced more ears m−2 than did euplasmic
‘Selkirk’. The increased tiller number and leaf area of
alloplasmic lines resulted in greater canopy light interception than euplasmic ‘Selkirk’ early in the season.
This characteristic was associated with reduced weed
populations under crops of alloplasmic ‘Selkirk’ lines
grown under low-, but not high-input, agronomic
regimes, with Ae. cylindrica– and Ae. ventricosa–
‘Selkirk’ significantly outyielding alloplasmic ‘Selkirk’
under low-input conditions. The F populations from
2
crosses between European wheat varieties and
‘Selkirk’ lines exhibited higher standard deviations for
grain yield for alloplasmic than for euplasmic ‘Selkirk’,
suggesting potential for selecting heterotic nuclear–
cytoplasmic combinations with alien cytoplasms.
Key words: Alien
photosynthesis.
cytoplasmic
variation,
wheat,
Introduction
In wheat (Triticum aestivum L.), alien cytoplasm substitution (resulting in alloplasmic lines) has been reported to
produce agronomically valuable changes in a range of
phenotypic traits, including plant height, flowering date
(Busch and Maan, 1978), cold resistance (Cahalan and
Law, 1979), disease resistance ( Washington and Maan,
1974), and disease tolerance ( Keane and Jones, 1990).
In contrast, the effects of alien cytoplasms on grain
yield have been reported to be largely detrimental
( Kihara, 1973; Law and Worland, 1984). The principal
trait affecting grain yield in wheat alloplasmics has been
cytoplasmic male-sterility, although certain alloplasmic
lines have been shown to exhibit potentially valuable
alterations in some of the components of economic yield,
such as ear number ( Fujigaki and Tsunewaki, 1979), dry
matter production (Hori and Tsunewaki, 1969), 1000
grain weight (Jost et al., 1975), and spikelet number per
ear (Netevic and Sanduhaz, 1968).
Most studies utilizing alien cytoplasms in wheat
improvement have centred on their use as inducers of
cytoplasmic male-sterility for F hybrid wheat breeding
1
programmes ( Wilson and Ross, 1962; Edwards, 1983;
Wilson and Driscoll, 1983). A number of researchers,
however, have investigated the possibility of using fertile
alloplasmic wheat lines to achieve nuclear–cytoplasmic
heterosis, with alloplasmic lines out-yielding the two
euplasmic parents (which contain nuclear and cytoplasmic
genomes from the same source). This has led to the
development of two contrasting strategies, with an
emphasis on either cytoplasms from species distantly
related to T. aestivum (to introduce a wider range of
effects; Yonezawa et al., 1986), or alternatively, on cytoplasms from species closely related to T. aestivum (to
minimize detrimental effects; Sasakuma and Ohtsuka,
1979). Results from this research have been equivocal.
Several groups, particularly those studying cytoplasms
from species closely related to T. aestivum, have reported
heterosis ( Kihara, 1973; Panayotov and Gotsov, 1973),
while others did not ( Edwards, 1983; Yonezawa et al.,
1986).
3 To whom correspondence should be addressed. Fax: +353 21 274420. E-mail: [email protected]
© Oxford University Press 1998
1520
Jones et al.
In plants, the cytoplasmic genomes, the chondriome
(containing approximately 120–140 genes; Sugiura, 1992)
in the mitochondria, and the plastome (95–100 genes;
Schuster and Brennicke, 1994) in the plastids, carry genes
which code predominantly for components of the gene
expression machinery (tRNA and rRNA) and for
membrane-bound proteins involved in organelle-specific
metabolism.
The transfer, during evolution, of many cytoplasmic
genes to the nuclear genomes has resulted in the dependence of mitochondria and plastids on the import of
nuclear-encoded proteins to carry out organelle biogenesis
and photosynthesis (chloroplasts) or respiration (mitochondria). This, in turn, necessitates modulation of cytoplasmic and nuclear gene expression, as illustrated by the
co-ordinated expression of the plastid-encoded rbc L gene
(coding for the large subunit) and the nuclear-encoded
rbc S gene (for the small subunit) responsible for Rubisco
synthesis. This inter-genomic regulation of gene expression is achieved by the involvement in the control of
cytoplasmic gene expression of nuclear-encoded gene
products including key structural and enzymatic factors
vital for organellar function, and regulatory factors, such
as those controlling plastid mRNA translation (Gillham
et al., 1994).
Given the involvement of both cytoplasmic and nuclear
genes in controlling carbon metabolism, the replacement
of parental chloroplast and mitochondrial genomes with
those from related species may be expected to impact on
photosynthesis and respiratory metabolism, either directly
or via modified nuclear–cytoplasmic interaction. Little
has been published in this area, although Evans (1986)
reported that an alloplasmic wheat line containing
Triticum boeoticum cytoplasm expressed only 71% of the
in vitro Rubisco activity exhibited by the euplasmic
(T. aestivum cytoplasm) line.
The objective of this research was to investigate the
effect of alien cytoplasms on carbon metabolism and
productivity of wheat in the glasshouse and in the field
under northern European conditions.
Materials and methods
Plant material
The cytoplasm donors of the alloplasmic series based on T.
aestivum cv. Chinese Spring ( Table 1) and T. aestivum cv. Chris,
cv. Selkirk and T. durum (Table 2) are given in the relevant
tables. These series were developed by Professor K Tsunewaki,
Kyoto University, Japan (‘Chinese Spring’ series) and Professor
SS Maan, North Dakota State University, USA (T. durum,
‘Chris’ and ‘Selkirk’ series) by crossing the euplasmic line (as
pollen parent) with the cytoplasmic donor (as egg parent) and
then back-crossing (generally for more than ten generations)
the hybrid (as female parent) to the recurrent euplasmic line.
Pot experiments
Plants were grown in a peat-based compost in 12.5 cm pots
(two plants per replicate pot, eight replicates per genotype) in
a glasshouse with supplementary lighting, giving maximum
irradiances of 880 mmol m−2 s−1 with a photoperiod of 16 h
and a temperature range of 18–25 °C. Plants were fed weekly
with half-strength Hoagland’s nutrient solution, with regular
flushing of the pots to prevent accumulation of salts.
Field experiments
Plants were grown in 1×1 m (F material ) or 2×2 m microplots
2
(other trials), at a plant density of 250 plants m−2, with four
replicates per treatment in a replicated randomized block
design. Fertilizer was applied to a total of 150 kg nitrogen ha−1,
in two splits (50 kg nitrogen ha−1 as 10:10:15 in the seed bed
and 100 kg nitrogen ha−1 as calcium ammonium nitrate at
GS30; Zadoks et al., 1974), unless stated to the contrary. To
prevent lodging, plants were supported by allowing them to
grow through 10×10 cm mesh netting. Fungicide and herbicide
applications were routinely made as used for the high-input
regime (described below), unless stated otherwise.
In the experiment comparing the field performance of
alloplasmic and euplasmic ‘Selkirk’ lines under high- and lowinput regimes, the former involved 200 kg nitrogen ha−1, a
three-spray fungicide programme (at first node (GS31), flag
leaf emergence (GS39) and anthesis (GS66; Zadoks et al.,
1974), using ‘Tilt C’ (Ciba-Geigy; active ingredients (a.i.)
propiconazole and carbendazim) at the manufacturer’s recommended rate, and a post-emergence herbicide treatment with
‘Ally’ (Du Pont; a.i. metsulphuron-methyl ) at the manufacturer’s recommended rate, at the fourth leaf stage. In the lowinput regime, the agrochemical treatments were adjusted to
80 kg nitrogen ha−1, a one-spray fungicide programme (at flag
leaf emergence) at the recommended rate, and a post-emergence
herbicide application made at one-quarter the recommended
rate. In both regimes, 50 kg nitrogen ha−1 was applied in the
seed bed, with the remainder at GS30. Weed biomass (aboveground tissue) was harvested from each microplot at crop
maturity, dried at 65 °C for 48 h and then weighed.
Measurements
Unless otherwise stated, all measurements on pot-grown plants
were conducted on tissues of the main stem. Measurements in
microplots were conducted on the whole plants (main stem plus
tillers). Weights were determined after plant material was dried
at 65 °C for 48 h.
Light interception measurements
In the study of four alloplasmics and the euplasmic ‘Selkirk’
line, light interception by the crop canopy was measured
between 12.00 h and 14.15 h at 15 dates throughout the growing
season. Six sites were selected at random at the base of each
microplot and the amount of photosynthetically active radiation
(PAR) was measured at each site on each date, using a PAR
sensor and meter (Skye Instruments, Llandrindod Wells, Powys,
UK ). These values were converted into percentage light
interception by reference to incident PAR measurements above
the canopy. Light interception during three phases of crop
development (GS1–GS30, GS31–GS65 and GS66–GS99) was
obtained by integration of the ‘% light interception versus time’
curve, using the trapezoid method, and expressed as Area
Under the Light Interception Curve (AULIC ).
Table 1. Morphological and developmental traits in the T. aestivum cv. Chinese Spring alloplasmic series
All traits except tiller number refer to the main shoot of the pot-grown plants. The absolute value is presented for euplasmic ‘Chinese Spring’ (T. aestivum cytoplasm); values for alloplasmic
lines are presented as % euplasmic value. An asterisk indicates a significant difference (P<0.05) from the corresponding euplasmic value, using the Protected Least Significant Difference,
following ANOVA of the absolute values.
Plasmatype
( Tsunewaki,
1988)
Plant
height
(cm)
Ear
length
(cm)
Spikelet
no.
Tiller
no.
Stem
diameter
(cm)
Grains
ear−1
Flowering
date (days
after June 1)
1000-grain
weight
(g)
Total
biomass
(g)
Grain
yield
(g)
Triticum aestivum
Aegilops ovata
Triticum boeoticum
Aegilops mutica
Aegilops umbellulata
Secale cereale
Aegilops bicornis
Aegilops squarrosa
Aegilops variabilis
S
M
A
Mt
U
R
Sb
D
S
84.4
99.0
84.6*
98.5
88.2*
97.3
90.1*
101.3
93.9*
5.6
105.7
107.7*
79.2*
106.7*
113.6*
110.4*
105.0
89.6*
16.9
104.3
94.7
90.6*
91.8*
101.0
99.7
98.8
99.5
2.7
81.2
113.0
126.8
118.8
130.6
108.7
140.6*
108.7
1.4
99.9
110.3
100.7
101.6
100.4
91.1
100.0
93.7
32.7
80.9*
48.6*
66.7*
17.3*
86.7*
95.1
107.6*
88.8*
20.0
232.5*
112.5
164.1*
130.0*
140.1*
110.6
103.5
104.4
30.6
73.8*
91.7
73.1*
87.5
80.2*
83.1*
82.6*
89.8
2.1
81.0
75.2
84.8
54.3*
80.2
85.2
93.3
91.4
1.0
54.8*
58.7*
68.3*
19.4*
70.7*
86.5
100.0
94.2
3.67
<0.001
16.63
<0.001
2.40
<0.05
2.55
<0.05
1.14
>0.05
12.34
<0.001
179.08
<0.001
7.60
<0.001
2.24
<0.05
89.4
<0.001
F
P
Photosynthesis in alloplasmic wheat
Cytoplasm donor
1521
1522
Jones et al.
Table 2. Plant height and flowering date in three alloplasmic wheat series
The values refer to the main shoot of pot-grown plants. An asterisk indicates a significant difference (P<0.05) from the corresponding euplasmic
value using Protected Least Significant Difference following ANOVA. na: alloplasmic line not available.
Cytoplasm donor
Ae. uniaristata
Haynaldia villosa
Ae. juvenalis
Ae. cylindrica
Ae. squarrosa
Ae. ventricosa
Ae. variabilis
Triticum macha-PI 140191
Triticum macha-PI 190923
T. turgidum
Euplasmic
F
P
Plasmatype
( Tsunewaki, 1988)
M
V
D
D
D
D
S
S
S
S
S
Height (cm)
Flowering date (days after 1 June)
‘Selkirk’
‘Chris’
T. durum
‘Selkirk’
‘Chris’
T. durum
80.4
77.5
80.0
78.5
81.0
80.0
77.5
77.6
82.5
80.0
80.6
88.1
60.9
88.8
97.5
95.3
97.0
81.8
na
na
na
89.8
59.8*
91.1
74.1*
87.5
73.5*
57.0*
84.1
na
na
na
87.4
21.6
20.8
20.4
20.8
21.2
21.5
22.3
21.4
20.1
20.0
20.9
25.1*
21.4
21.5
24.5*
23.0*
23.0*
29.8*
na
na
na
21.4
36.4
36.5
29.4*
44.8*
40.4
56.0*
38.3
na
na
na
40.1
0.83
>0.05
2.24
<0.05
12.58
<0.001
2.51
<0.05
6.70
<0.001
14.73
<0.001
Photosynthesis measurements
Photosynthesis was measured on main shoot flag leaves of
pot-grown plants of the ‘Selkirk’ alloplasmic series at
anthesis. Photosynthetic rates were measured using an ADC
portable gas-exchange system and data logger (LCA2;
Analytical Development Company, Hoddesdon, UK ), during
10.00–16.00 h. Leaves were illuminated with a Hansatech light
source (LS2H; Hansatech Ltd, King’s Lynn, UK ), containing
appropriate heat and neutral density filters (Balzers,
Liechtenstein). The lamp was positioned to provide an even
illumination of the leaf chamber window. Leaf temperature was
maintained at 20 °C with a vapour pressure deficit of ~0.8 kPa.
Prior to the determination of P , the dark respiration
max
rate was measured and the leaf then subjected (for 5 min)
to two intermediate and progressively higher light levels,
before measuring the photosynthetic rate after it had
stabilized (10–15 min) at the saturating irradiance (1920 mmol
photon m−2s−1). This protocol was used to minimize the
possibility of photoinhibition. The irradiance incident on the
leaf tissue was measured with a Skye quantum sensor and meter
(Skye Instruments, Llandrindod Wells, UK ). Reduced nitrogen
in the flag leaf segment used for photosynthetic studies was
determined using the semi-micro Kjeldahl method. Flag leaf
area was measured using a Li-Cor model LA–3000 leaf area
meter (Li-Cor, Nebraska, USA). Total chlorophyll, chlorophyll
a and chlorophyll b measurements were carried out using the
method of Harborne (1984).
Statistical analysis
Date analysis was conducted using parametric ANOVA.
Comparisons between the euplasmic line and individual alloplasmics were conducted using the Protected Least Significant
Difference (P<0.05), i.e. comparisons were conducted only where
the sample F value (sample mean square divided by residual
mean square) was significant (P<0.05). Comparison of euplasmic
and mean alloplasmic values were conducted using t-tests.
Results
Alloplasmic-euplasmic comparison of pot-grown plants
Initial studies were made on pot-grown plants of the T.
aestivum ‘Chinese Spring’ alloplasmic series which is the
largest available for any nuclear parent, covering a wide
range of plasmatypes. Of the ten traits investigated, only
stem diameter was not significantly affected by cytoplasmic substitution ( Table 1), with the Ae. squarrosa
cytoplasm affecting the fewest characters.
Several alloplasmic lines of ‘Chinese Spring’ exhibited
lower grain yield than euplasmic ‘Chinese Spring’; this
was due largely to partial male sterility, manifested as
fewer grains per ear. These lines included those with
Ae. mutica, T. boeoticum or Ae. umbellulata cytoplasm
( Table 1). Only plasmatype S (Ae. bicornis, Ae. variabilis)
and plasmatype D cytoplasms (Ae. squarrosa) resulted in
yields which were not significantly lower than that of
euplasmic ‘Chinese Spring’.
A narrower range of cytoplasms, concentrating on
plasmatypes S and D, was then studied, using alloplasmic
series based on T. aestivum cvs ‘Selkirk’ and ‘Chris’ and
T. durum ( Tables 2, 3). Of these, ‘Chris’ and ‘Selkirk’
appeared to be less affected by cytoplasmic substitution
than T. durum, with ‘Selkirk’ being the least affected. The
effect of individual cytoplasms on relatively simplyinherited traits such as flowering date, flag leaf area and
tiller number appeared to be more similar when two
aestivum parents were being compared, than when aestivum and durum were studied. For flowering date ( Table 2)
for example, correlation coefficients of r=+0.58
(‘Selkirk’ versus ‘Chris’), r=+0.27 ( ‘Selkirk’ versus
durum) and r=+0.01 (‘Chris’ versus durum) were
obtained although none of these coefficients were statistically significant. No trends were evident for more complex
traits; for grain yield ( Table 3); for example, correlation
coefficients ranged from r=+0.17 (‘Selkirk versus ‘Chris)
to +0.18 (‘Selkirk’ versus durum) and −0.23 (‘Chris’
versus durum).
The wide variation in grain yield in the T. durum series
( Table 3) was associated with reduced male fertility in
Photosynthesis in alloplasmic wheat
1523
Table 3. Grain yield and grain number per ear in three alloplasmic wheat series
The values refer to the main shoot of pot-grown plants. An asterisk indicates a significant difference (P<0.05) from the corresponding euplasmic
value, using the Protected Least Significant Difference following ANOVA. na: alloplasmic line not available.
Cytoplasmic donor
Ae. uniaristata
H. villosa
Ae. juvenalis
Ae. cylindrica
Ae. squarrosa
Ae. ventricosa
Ae. variabilis
T. macha-PI 140191
T. macha-PI 190923
T. turgidum
Euplasmic
F
P
Plasmatype
( Tsunewaki, 1988)
M
V
D
D
D
D
S
S
S
S
S
Grain yield (g) per plant
Grain number per ear
‘Selkirk’
‘Chris’
T. durum
‘Selkirk’
‘Chris’
T. durum
0.86
0.81
0.84
0.96
0.90
0.84
0.83
0.94
0.94
0.98
0.92
0.58
0.58
0.83
0.74
0.93
0.82
0.41
na
na
na
0.77
0.11*
0.80
0.17*
0.52*
0.39*
0.12*
0.75
na
na
na
0.73
22.8
20.5*
21.7*
23.4
23.0
24.2
19.9*
20.8*
26.1
26.2
25.2
23.9*
32.0
34.1
38.9
40.5*
34.9
21.6*
na
na
na
34.0
9.8*
35.3
7.1*
27.9
18.1*
6.4*
32.6
na
na
na
31.0
0.73
>0.05
3.45
<0.01
4.31
<0.001
2.73
<0.05
5.46
<0.001
20.3
<0.001
lines with cytoplasm from Ae. juvenalis, Ae. ventricosa,
Ae. squarrosa (all plasmatype D) and Ae. uniaristata
(plasmatype M ). Partial male sterility was much less
evident in the alloplasmic lines of the T. aestivum cultivars,
with only Ae. variabilis and Ae. uniaristata expressing
reduced seed set, and then only when in combination
with the ‘Chris’ nuclear genome ( Table 3).
The alien cytoplasms studied here proved to have less
detrimental effects when in combination with an aestivum
nuclear genome than with the durum genome. Within
the aestivum alloplasmic series, alloplasmics with D
plasmatype cytoplasms consistently resulted in yields similar to that of the corresponding euplasmic line ( Table 2),
as did those with the S plasmatype cytoplasms (except
for the Ae. variabilis–‘Chris’ combination which exhibited
partial male sterility).
Alloplasmic–euplasmic comparison of field-grown crops
No significant difference was observed between the yields
of seven selected D and S plasmatype alloplasmic lines
and euplasmic ‘Selkirk’ (Table 4). The only statisticallysignificant differences observed were for ear number m−2,
with Ae. ventricosa ‘Selkirk’ having a higher ear
number m−2 than euplasmic ‘Selkirk’ and several of
the S plasmatype ‘Selkirk’ alloplasmics ( Table 4).
Comparison of the mean values for lines of the two
plasmatypes identified inter-plasmon differences for the
characters ear number m−2 (mean of 407.3 for D cytoplasms, 371.3 for S cytoplasms (including euplasmic
‘Selkirk’); t=5.21, P<0.01) and grain number ear−1 (31.0
for D, 33.1 for S; t=2.63, P<0.05), but not for
1000-grain weight (41.5 g for D, 40.0 g for S; t=1.96,
P>0.05) or grain yield (0.52 kg m−2 for D, 0.49 kg m−2
for S; t=1.14, P>0.05).
The increase in ear number m−2 associated with D
cytoplasm alloplasmics ( Table 4) was investigated further.
A significant positive correlation (r=+0.89, n=8;
P<0.05) was observed between ear number m−2 and
grain yield for the eight ‘Selkirk’ lines ( Table 4), although
the partial autocorrelative nature of this association
should be noted. By GS30, field-grown D plasmatype
alloplasmic lines had produced significantly more tillers
than euplasmic ‘Selkirk’, as had two of the S cytoplasmics,
T. macha–‘Selkirk’ and T. turgidum–‘Selkirk’, although
the differences in numbers of tillers surviving to maturity
were not significant ( Table 5). When the height of the
first-formed tiller was expressed as a percentage of that
of the main shoot, it was evident that there was a greater
difference in height of the two earliest-produced stems in
euplasmic ‘Selkirk’ than in the S cytoplasm alloplasmics
(non-significant) and the D cytoplasm alloplasmics
(P<0.05) ( Table 5). These data suggest that apical dominance was reduced in the alloplasmic lines, compared to
euplasmic ‘Selkirk’.
Photosynthesis measurements
The photosynthetic capacity of the main shoot flag leaf
was compared at anthesis in glasshouse-grown plants of
alloplasmic (D and S cytoplasmics) and euplasmic
‘Selkirk’ lines. No significant differences in P
per unit
max
leaf area or in flag leaf area were observed, although the
area of the flag leaf of each of the alloplasmic lines,
particularly those carrying S plasmatype cytoplasms, was
larger than that of euplasmic ‘Selkirk’. Similar trends
were observed when total photosynthetic activity of the
flag leaves (i.e. P ×leaf area) was compared, with the
max
three S cytoplasm alloplasmic lines exhibiting the highest
values. Although no significant inter-line differences were
detected for these characters, significant differences
occurred between the mean alloplasmic and the euplasmic
value for both flag leaf area (0.00327 m2 and 0.00270 m2,
respectively; t=2.53, P<0.05) and total flag leaf
1524
Jones et al.
Table 4. Grain yield and yield components from microplots of the T. aestivum cv. Selkirk alloplasmic series
The values refer to the total from main stems plus tillers of the field-grown plants. An asterisk indicates a significant difference (P<0.05) from the
euplasmic value (T. aestivum cytoplasm), using the Protected Least Significant Difference, following ANOVA.
Cytoplasm donor
Plasmatype
( Tsunewaki, 1988)
Grain yield
(kg m−2)
Ears m−2
Grains ear−1
1000-grain
weight (g)
Total biomass
(kg m−2)
Harvest
index (%)
Ae. juvenalis
Ae. cylindrica
Ae. ventricosa
Ae. squarrosa
Ae. variabilis
T. turgidum
T. macha PI 190923
T. aestivum
D
D
D
D
S
S
S
S
0.49
0.51
0.56
0.53
0.44
0.54
0.48
0.50
402
395
423*
409
348
413
355
369
29.7
31.9
31.3
30.9
33.1
32.9
33.3
33.1
41.1
40.5
42.3
42.0
38.6
39.8
40.6
41.0
1.30
1.24
1.43
1.46
1.20
1.35
1.25
1.39
37.7
41.2*
39.2
36.3
36.6
40.1
38.4
36.1
1.57
>0.05
4.01
<0.01
2.13
>0.05
2.21
<0.05
2.06
>0.05
2.51
<0.05
F
P
Table 5. Tiller number and apical dominance in field-grown plants
of the alloplasmic series of T. aestivum cv. Selkirk
Measurements were made on field-grown plants, at GS (growth stage)
values according to Zadoks et al. (1974). Apical dominance (%) was
expressed as (height of first tiller)×100[height of main shoot. An
asterisk indicates a significant difference (P<0.05) from the euplasmic
values (T. aestivum cytoplasm), using the Protected Least Significant
Difference, following ANOVA.
Cytoplasmic parent
(plasmatype)
Tiller number per plant
Apical
dominance (%)
GS 30
GS 99
GS 99
Ae. juvenalis (D)
Ae. cylindrica (D)
Ae. ventricosa (D)
Ae. squarrosa (D)
Ae. variabilis (S)
T. turgidum (S )
T. macha PI 190923 (S)
T. aestivum (S)
5.40*
5.31*
5.61*
5.06*
4.51
5.31*
5.16*
4.37
3.10
3.38
3.41
3.27
3.19
3.04
2.85
2.91
91.6*
89.5
92.3*
90.9*
87.6
85.9
88.6
83.8
F
P
3.91
<0.01
2.01
>0.05
3.61
<0.01
photosynthesis activity (0.0436 mmol CO leaf −1 s−1
2
and 0.355 mmol CO leaf −1 s−1, respectively; t=2.83,
2
P<0.05). No significant difference was observed in P
max
between the mean alloplasmic (13.3 mmol CO m−2 s−1)
2
and the euplasmic values (13.4 mmol CO m−2 s−1)
2
(t=0.43, P>0.05). No significant correlation was
observed between P
(expressed on a leaf area basis)
max
and flag leaf area (r=−0.22, n=8). On the other hand,
significant correlations were obtained between P
max
and the flag leaf traits chlorophyll content (r=+0.76,
n=8; P<0.05), chlorophyll a:b ratio (r= −0.86,
n=8; P<0.05), chlorophyll b content (r=+0.85, n=8;
P<0.01) and nitrogen content (r=+0.90, n=8,
P<0.01).
Light interception studies
The impact on canopy light interception of the higher
tiller production and larger flag leaf areas of selected D
and S cytoplasm alloplasmic lines was investigated in
field microplots. Seasonal light interception (expressed as
Area Under the Light Interception Curve, AULIC ) by
the four alloplasmic lines, especially those with D cytoplasms, was greater than that of euplasmic ‘Selkirk’
( Table 6). The seasonal AULIC was then split into three
important phases of crop development: GS1–GS30 (tillering), GS31–GS65 (stem extension), and GS66–GS99
(post-anthesis). The general trend was similar for all four
alloplasmics. Light interception by each alloplasmic line
was higher than that exhibited by the euplasmic line
in the first two phases, but was significantly lower than
that of the euplasmic during the post-anthesis period
( Table 6). The lower light interception late in the season
was associated with more rapid leaf senescence in the
alloplasmic lines.
Productivity under high- and low-input regimes
The field performances of the four alloplasmic lines and
euplasmic ‘Selkirk’ were compared under both high-input
(with respect to use of agrochemicals such as fertilizers,
fungicides and herbicides) and low-input conditions. No
significant yield differences were observed under the highinput regime, but under the low-input conditions, all four
alloplasmics produced higher yields than did euplasmic
‘Selkirk’. The yields of the D cytoplasm alloplasmics were
significantly higher than that of the euplasmic line
( Table 7). Under the low-input regime, the alloplasmic
crops also supported a smaller weed population than the
euplasmic ‘Selkirk’ ( Table 7).
Modification of nuclear genotype in alloplasmic lines
In the previous experiments, the T. aestivum parental
varieties were spring wheat varieties (‘Selkirk’ and ‘Chris’)
adapted to North American conditions. To determine
whether substitution of D and S cytoplasms would have
similar effects on European spring wheat varieties, alloplasmic and euplasmic ‘Selkirk’ were crossed with two
Photosynthesis in alloplasmic wheat
1525
Table 6. Canopy light interception (AULIC) on microplot trials of lines of the T. aestivum cv. Selkirk alloplasmic series
Light interception was determined as Area Under the Light Interception Curve (AULIC ). An asterisk indicates a significant difference (P<0.05)
from the euplasmic value, using the Protected Least Significant Difference, following ANOVA.
Cytoplasm donor
Ae. cylindrica
Ae. ventricosa
T. macha PI 190923
T. turgidum
T. aestivum
Plasmatype
( Tsunewaki 1988)
D
D
S
S
S
F
P
AULIC
GS1–GS30
GS31–GS65
GS66–GS99
Total (GS1–GS99)
8.4*
8.9*
7.0
8.3*
5.7
23.8*
25.0*
24.1*
23.3*
19.5
42.8
40.7*
40.7*
38.8*
45.1
75.0*
74.6*
71.8
70.4
70.3
4.97
<0.05
4.89
<0.05
6.31
<0.01
4.80
<0.05
Table 7. Grain yield of lines of the T. aestivum cv. Selkirk alloplasmic series in microplot trials under high- and low-input regimes
Microplots (2×2 m) of wheat were grown under high- (200 kg nitrogen ha−1, 3-spray fungicide programme, full-rate post-emergence herbicide
spray) and low-input regimes (80 kg nitrogen ha−1, 1-spray fungicide programme, one-quarter full-rate post-emergence herbicide spray). An asterisk
indicates a significant difference (P<0.05) from the euplasmic value (T. aestivum cytoplasm), using the Protected Least Significant Difference,
following ANOVA.
Cytoplasm donor
T. aestivum
Ae. cylindrica
Ae. ventricosa
T. macha PI 190923
T. turgidum
Plasmatype
(Tsunewaki,
1988)
S
D
D
S
S
F
P
High-input regime
Low-input regime
Grain yield
(g m−2)
Grain yield
(g m−2)
Weed biomass
(g m−2)
368.4
360.7
384.6
347.3
381.5
242.3
290.6*
281.5*
261.3
267.3
78.4
47.8*
45.9*
45.1*
70.1
2.89
>0.05
3.72
<0.05
8.93
<0.001
commercial European-bred spring wheats, ‘Alexandria’
and ‘William’. No statistically-significant differences were
observed between the yields of the F hybrids with
1
aestivum or alien cytoplasm ( Table 8), although the D
cytoplasm hybrids consistently produced higher grain
Table 8. Grain yields of F and F populations from crosses
1
2
between Selkirk lines (euplasmic and alloplasmic) and two spring
wheat varieties
Grain yield per plant (main stem and tillers) was determined on potgrown (F generation) and field-grown plants ( F generation).
1
2
Nuclear hybrid
Cytoplasm donor
(plasmatype
Grain yield
(g per plant)
F
1
F (±SD)
2
‘Selkirk’בAlexandria’ Ae. cylindrica (D)
Ae. ventricosa (D)
T. turgidum (S)
T. aestivum (S )
‘Selkirk’בWilliam’
Ae. cylindrica (D)
Ae. ventricosa (D)
T. turgidum (S)
T. aestivum (S )
2.51
2.62
2.45
2.34
2.85
2.80
2.60
2.69
2.15±0.52
2.30±0.60
2.07±0.53
2.14±0.41
2.56±0.70
2.46±0.62
2.38±0.58
2.35±0.44
F
P
1.21
>0.05
1.59
>0.05
yields than the euplasmic hybrids. Similar results were
obtained from the F generations. Of particular interest
2
was the observation that the F populations with alien
2
cytoplasms exhibited higher standard deviations for grain
yield than did the corresponding euplasmic F populations
2
( Table 8).
Discussion
The multiple backcross generations employed in the development of the alloplasmic lines should have minimized
the retention of alien nuclear genes from the egg parent
in the alloplasmic lines. Phenotypic differences between
the corresponding euplasmic and alloplasmic lines should
thus be largely attributable to differences in content of
the cytoplasmic genomes.
The results from the ‘Chinese Spring’ alloplasmic series
support the plasmatype characterization of Tsunewaki
(1988); only the D and S plasmatype cytoplasm (classified
by Tsunewaki et al. (1983) as having the least effects on
plant fertility and growth, as a result of being closely
related to T. aestivum) produced alloplasmic lines with
yields similar to that of the euplasmic line. The good
agronomic performances of D and S cytoplasm alloplas-
1526
Jones et al.
mics reported here also agree with results from Japan
(Sasakuma and Ohtsuka, 1979) and North America
(Busch and Maan, 1978). Some differences in responses
of individual alloplasmics compared to studies published
previously were noted and these may reflect environmental effects on nuclear–cytoplasmic interactions. For
example, Ae. bicornis cytoplasm caused delayed heading
of ‘Chinese Spring’ in these studies, but not in similar
work carried out in Bulgaria (Panayotov and Gotsov,
1976).
The effect of cytoplasm substitution independent of the
nuclear (recurrent) parent on relatively simply inherited
characters, such as heading date, supports earlier reports
from this laboratory on traits such as tolerance of, and
resistance to, Stagonospora (Septoria) nodorum infection
( Keane and Jones, 1990).
Different S plasmatype cytoplasms resulted in different
phenotypic effects on the same nuclear parent (primarily
‘Selkirk’ in this study). Of the four alien S cytoplasms,
turgidum–‘Selkirk’ and both macha–‘Selkirk’ lines were
superior in agronomic terms to ‘Selkirk’ with aestivum
cytoplasm (another S plasmatype cytoplasm). Ae. variabilis cytoplasm was generally the poorest of the S
plasmatype cytoplasms in combination with the ‘Selkirk’
nuclear genome, a finding also reported from North
American studies ( Kofoid and Maan, 1982). The two
T. macha accessions also produced different effects.
Generally, the alloplasmic carrying the cytoplasm from
T. macha accession PI 190923 proved to have a superior
field performance, and this line was used in subsequent
investigations.
Among the D plasmatype cytoplasms used, Ae. squarrosa is considered to be the cytoplasmic donor for both
Ae. cylindrica and A. ventricosa (Maan, 1978), so the
similar effects observed for alloplasmics carrying cytoplasm from these three species is not surprising. Microplot
studies on the selected male-fertile D and S cytoplasm
‘Selkirk’ alloplasmics showed that they exhibited
increased tiller production, compared to euplasmic
‘Selkirk’. This appeared to be associated with reduced
apical dominance in the alloplasmic lines, the height of
the tillers being closer to that of the main shoot in this
material than in euplasmic ‘Selkirk’. Increased tillering
may have contributed to the high yields exhibited in the
field by these alloplasmic lines, either as a direct result of
increased density of ears in the population (an important
component of grain yield ), or by increasing light interception up to anthesis ( Table 6).
Despite the importance of plastome genes in chloroplast
metabolism, no consistent effects of alien cytoplasm substitution on P
were observed (Table 6). The small
max
changes in P
were paralleled by changes in stomatal
max
conductance and, therefore, there was little variation in
intercellular CO concentration. Evans (1986) reported
2
that ‘Chinese Spring’ with T. boeoticum cytoplasm had
decreased Rubisco activity in vitro, although this was not
translated into an altered net photosynthesis rate in vivo.
Between the near-isogenic lines of the ‘Selkirk’ alloplasmic series, photosynthesis rate per unit leaf area was
positively correlated with chlorophyll content, in particular chlorophyll b content, leading to a negative correlation
with chlorophyll a5b ratio. A similar set of associations
was reported by Watanabe et al. (1994) in a study of
twentieth century Australian wheat varieties. As chlorophyll b is exclusively located in the light-harvesting antennae, the correlation with chlorophyll b content suggests
that the variation in P
between lines of the ‘Selkirk’
max
alloplasmic series was more closely associated with differences in light-harvesting than with biochemical electron
transport processes. It is uncertain, however, how variations in light harvesting could affect the maximum
photosynthetic capacity. The highly significant positive
correlations between P
and leaf nitrogen content (and
max
chlorophyll content) might suggest that the lines differed
in the ‘availability of nitrogen to the plants’. Trials on
the alloplasmic lines indicated that higher nitrogen fertilizer amounts did not significantly affect plant growth and
productivity (data not shown), indicating that nitrogen
was not growth limiting in these experiments. It is considered more likely that the lines differed in their ability to
partition nitrogen into leaf components which determine
P . These are thought to be primarily Rubisco, although
max
components of electron transport pathways could also be
involved as these also comprise a significant proportion
of the total leaf nitrogen ( Evans, 1989).
Whereas cytoplasm substitution appeared to have no
marked effect on the photosynthetic processes, it resulted
in a cytoplasm-independent increase in flag leaf area in
the alloplasmic lines, compared to euplasmic ‘Selkirk’.
Many published studies have reported negative correlations between wheat leaf area and photosynthesis rate
per unit leaf area (Austin et al., 1982; Bhagsari and
Brown, 1986; Lawlor, 1995), but the cytoplasm-associated
increase in flag leaf area caused no concomitant reduction
in photosynthesis rate per unit leaf area. This phenomenon opens up the (albeit distant) prospect of increasing
net assimilation rate per leaf and consequently crop
canopy net assimilation rate by increasing leaf area as
the result of alien cytoplasm substitution.
One detrimental trait exhibited by D and S cytoplasm
‘Selkirk’ alloplasmics was an increased rate of leaf senescence post-anthesis, and hence a reduced duration for the
grain-filling period. A similar trait has been reported for
primitive wheats in a comparison with modern wheat
cultivars ( Evans, 1993).
The increased light interception by alloplasmic canopies
early in the season ( Table 6), associated with the production of more tillers and larger leaves, suggested a potential
for greater competitiveness with weeds. Combined with
the increased yield tolerance to S. nodorum exhibited by
Photosynthesis in alloplasmic wheat
‘Selkirk’ alloplasmics ( Keane and Jones, 1990), these
lines may be particularly useful under low-input agricultural regimes. Microplot trials confirmed the hypothesis
of higher yield and increased weed control exhibited by
the alloplasmics (Table 7), although the relative contributions of increased assimilation, improved weed control
(both resulting presumably from greater light interception
by the crop canopy), and increased Stagonospora tolerance (S. nodorum and Erysiphe graminis f. sp. tritici, were
the major pathogens of the low-input microplots) were
not determined.
The large number of phenotypic traits influenced by
cytoplasmic substitution seems out of proportion to the
small number of different cytoplasmic genes, representing
less than 1% of the genes in the nuclear genome. The
highly conserved nature of many of the cytoplasmic genes,
e.g. tRNA and rRNA genes, argues against the hypothesis
that alien plastids or mitochondria could contain sufficiently high levels of genetic variation in transcribed
sequences to cause the observed phenotypic effects. A
more feasible scenario would involve differences between
the alien and recurrent parents in regulatory sequences in
cytoplasmic genes, so that nuclear–cytoplasmic interactions (‘cross-talk’) are disturbed in alloplasmics, compared to the corresponding euplasmics, resulting in
phenotypic variation.
To determine whether substitution of alien cytoplasm
(particularly of the D plasmatype) could be of potential
benefit in European wheat breeding programmes, the
cytoplasms would need to be combined with locallyadapted nuclear genomes. Studies on the F generation
2
of a crossing programme between alloplasmic and
euplasmic ‘Selkirk’ lines and two European spring wheat
varieties ( Table 8) suggested that small yield improvements could be achieved. The increased variation exhibited within alloplasmic, compared to euplasmic F
2
populations, suggests that selection for increased nuclear–
cytoplasmic heterosis could be made within segregating
populations.
Several authors in the 1980s suggested that alien cytoplasm substitution represented a potential strategy for
wheat improvement ( Yonezawa et al., 1986), but little
progress has been made in the past decade. Cytoplasm
substitution can result in the modification of a number
of plant traits, due to changes in chloroplast and mitochondrial genomes and alterations in nuclear–cytoplasmic
interactions. In addition to the beneficial effects described
here and by other authors, detrimental phenotypic
changes have also been attributed to alien cytoplasm
substitution, including premature grain sprouting
( Tsunewaki et al., 1983) and low-temperature variegation
(Cahalan and Law, 1979), although these effects was not
observed in the current study. The use of D plasmatype
cytoplasms would be expected to minimize the introduction of such characters (Panayotov, 1983). Indeed, Ae.
1527
ventricosa cytoplasm, possible the most useful alien cytoplasm identified in this study, has already been used
in breeding two commercial French wheat varieties,
‘Rendezvous’ and ‘Roazon’.
Acknowledgements
The authors would like to thank Professor K Tsunewaki,
Professor SS Maan and Professor C Konzak for their generous
gifts of seeds of the wheat lines used in this study.
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