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Conversely, mature embryos are available without limit at any time,
but are the least frequently used as an explant source because of the
low frequency of callus induction. However, recently developed
techniques (Bartok and Sagi 1990; Ahmed et al. 1992; Ozgen et al.
1996) such as endosperm- supported mature embryo methods have been
approved to induce callus quicker and with higher regeneration
capacity depending on the genotypes (Ozgen et al.1996, 1998). As a
known fact, callus induction and regeneration is influenced by the
genotype (Sears and Deckard 1982; Mathias and Simpson 1986; Chowdhury
et al. 1991), source of explant (Ozias-Akins and Vasil 1982; Redway
et al.1990) and culture media (Mathias and Simpson 1986; Fennel et
al. 1996). Besides, in endosperm- supported mature embryo culture,
the size of the seed and consequently the size of the endosperm may
additionally influence the callus induction and plant regeneration,
since calli use the metabolites of the endosperm.
The objective of this study was to reveal the correlation between the
different seed sizes and responses of mature embryo culture, in
different wheat (Triticum spp.) genotypes using
endosperm-supported mature embryo culture method.
Materials and methods
Four species of three ploidy groups of wheat; diploid (T.
monococcum L. and T. boeoticum L.), tetraploid
(T. durum Desf. 'Kunduru-1149') and hexaploid (T.
aestivum L. 'BezostaJa-1') were used as sources of mature
embryos. For each genotype, seed samples were grouped as large and
small after they were sieved.
Seeds were surface sterilized for 5 min in 70% (v/v) ethanol, rinsed
two times with sterile distilled water and followed by 25 min
incubation in a commercial sodium hypochlorite solution containing a
few drops of Tween 20. Finally, they were rinsed thoroughly at least
7 times with sterile distilled water. The seeds were then imbibed in
sterile water for 11 h at 26C. For hulled wheats, surface
sterilization and imbibition procedures were applied after the hulls
are removed.
For callus induction, mature embryos were aseptically moved slightly
with a scalpel but not excised and were placed furrow downwards in
sterile 10 cm petri dishes containing 8 mg/l 2,4-D, pH 5.8 solution
which was autoclaved at 121C for 15 minutes. Afterwards, they were
left for incubation at 26C in the dark for 11 days. At the end of the
incubation period, the number of calli and their fresh weights were
calculated.
The calli were transferred into hormone-free MS medium (Murashige and
Skoog 1962) containing (2mg/l) glycine, (20g/l) sucrose and (7g/l)
agar at pH 5.8. The transferred callus cultures were incubated at 26C
in the dark for 3 weeks and grown further at 26C 16 h light and 8 h
dark period in a fresh medium prepared in the same way for
regeneration. Photoperiod was obtained by using 40W tungsram (1500
lux) fluorescence lights.
A completely randomized design with three replications per seed group
for each genotype was used. Petri dishes containing 20 seeds were
considered the units of replication (total 120 seeds per genotype).
The effect of species on culture responses was determined by analysis
of variance and least significant difference (LSD) tests..
Correlation coefficients were calculated for relationships between
different characters. Differences between large and small seeds were
evaluated with the chi-square test for independence (Steele and
Torrie 1960).
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