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Materials and methods
Plant materials and growth condition
Six American triticale lines; 6A-845, 6A-854, 6A-1092 (ROSNER++),
6A-1093 (ROSNER--), 801-1210 and Polish line 'Salvo' were used in
this study. Plants were grown in 5-liter pots containing a mixture of
vermiculate soil and chemical fertilizer (N:P:K=1:1:1). All plants
were established outdoors under natural photoperiod with day/night
temperature of 30/20C and an average photosynthetic photon flux
density of 1600 microE
m-2s-1.
All plants were daily watered to avoid any water stress effects.
Fully expanded leaves from five week-old plants were used in the
experiments. Five independent measurements per accession were made,
and the mean values and standard errors were calculated.
Photosynthesis
Leaf gas exchange rates were monitored with single attached leaf
enclosed in an acrylic plastic leaf chamber (6x10x0.5 cm). 1 KW metal
halide lamp provided the light source (1500
microEm-2s-1).
The air stream (with 340 microLL-1 CO2) was
passed into the leaf chamber at a constant rate of 3L
min-1. The CO2 uptake was measured with an
infrared gas analyzer (Analytical Development Co., Hoddesdon U.K.).
Photon flux at the surface of each leaf was measured with a quantum
sensor (Li-Cor Corporation, Nebraska, U.S.A.). Leaf temperature was
measured with finewire thermocouples pressed to the lower surface of
the leaf and was maintained at 25 + or - 2C by adjusting the
temperature of the chamber by running water through water jacket.
Enzyme assay and kinetic properties of RuBP carboxylase
The activity of RuBP carboxylase, its kinetic characteristics and
total chlorophyll content were determined on the same leaves used for
the gas exchange studies. The leaves (2g) were washed quickly, dried
and homogenized in 10ml of 100mM HEPES buffer (pH 7.8) containing 5mM
DTT, 1 mM EDTA, 25 mM MgCl2 in a chilled mortar. The
homogenate was centrifuged at 30,000 xg for 20 min at 0-3C. The
supernatant was passed through Sephadex G-200 which was equilibrated
with 100 mM HEPES buffer containing 20 mM MgCl2 and 10 mM
NaHCO3. The eluates were collected and assayed at 25C for
enzyme activity. The reaction mixture contained 50 mM HEPES-NaOH
buffer (pH 8.0), 5mM DTT, 20 mM MgCl2, 0.5 mM
NaH14CO3, 0.5 mM RuBP and the enzyme extract.
The acid stable radioactivity was determined in Beckman LS 1800
liquid scintillation system (Ramachandra Reddy and Das 1986).
Chlorophyll was estimated according to Arnon (1949).
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