Joint International Conference on Long-term Experiments ...

MATERIALS AND METHODS
Shoot cultures of eight pea genotypes (Graphis, Baccara, 2107, Hanka, Hunor, Janus,
M49060 and Erbi) were maintained in vitro and explants for the experiments were
isolated from micropropagated plants. Two kinds of explants were used: fully
developed leaflets of the upper third of stem or stem segments without node. Isolation
was made in a sterile solution (0.15 mgl -1 citric acid and 0.1 mgl -1 ascorbic acid) to
prevent the withering of explants and the deleterious oxygenation process. Explants
were placed on G 57 media (salts and vitamins) (Gamborg et al., 1968) supplemented
with MS-Fe (Murashige & Skoog, 1962), 3.0 % sucrose, 0.8 % agar-agar and 1.0 mgl -1
benzyl-adenine (BA) as cytokinin. Naphthalane-acetic acid (NAA), indolebutyric acid
(IBA) and 2,4-dichlorophenoxyacetic acid (2,4-D) were applied as auxins in two
concentrations: 2.0 and 4.0 mgl -1 . Cultures were incubated in dark for 3 weeks at 26 ºC,
after then they were cultured on 16 h photoperiod for further 4 weeks. Callus forming
capacity was observed and scored weekly as follows:
1- there is no callus on the explant
2- some calluses on some end of explant
3- there are some calluses on both end of explant
4- strong callus forming but the surface of the explant can be seen
5- the callus overgrows the explant.
Statistical analyses were made by variance analyses followed by Tukey test, using by
SPSS 9.0 for Windows programme.
RESULTS AND DISCUSSION
In our experiments callus induction and development were obtained in all treatments
and for each genotype. The statistical analysis showed significant interactions between
genotypes, explant type and auxins. In the average of all treatments the IBA was the
best auxin, and the 2,4-D was the least efficient. Effect of NAA in lower concentration
(2.0 mgl -1 ) was similar to the effect of IBA, although by the end of fourth week the
callus forming capacity was significantly lower (table 1.).
Table 1. The main-effect of auxin on callus forming capacity (in the average of both
explants and genotypes).
Callus development on the end of
1 st 2 nd 3 rd 4 th
Auxin
treatments
mgl -1 week
NAA 2.0 1.69 d 2.52 d 3.40 cd 4.25 cd
NAA 4.0 1.59 c 2.43 c 3.34 c 4.21 c
2,4-D 2.0 1.51 b 2.22 b 2.81 b 3.39 b
2,4-D 4.0 1.34 a 2.01 a 2.63 a 3.21 a
IBA 2.0 1.64 d 2.52 d 3.47 d 4.36 e
IBA 4.0 1.64 d 2.54 d 3.47 d 4.32 de
Means within column signed by the same letter belong to the same homogeneous group.
197