INFLUENCE OF PLASMOGENES ON THE PRODUCTIVITY OF MORPHOGENESIS IN STRAWBERRY ( FRAGARIA X ANANASSA DUCH . )

Plasmogenes are largely located in mitochondria or plastids and they can infl uence the inheritance of many plant characteristics. This phenomenon is called cytoplasmic inheritance and can be detected on the basis of the expression of a trait in progeny F1 obtained from single and reciprocal crosses. The aim of this study was to examine the cytoplasmic inheritance of in vitro productivity of morphogenesis in three genotypes of Fragaria x ananassa Duch., i.e. the cultivars ‘Dukat’, ‘Teresa’ and the breeding clone no. 590. Single and reciprocal crosses were done according to Griffi ng’s method 3. The value of general combining ability (GCA) indicated cv. ‘Teresa’ as the best maternal component for crossing and ‘Dukat’ as the worst. The negative reciprocal cross effects (rij) revealed the cytoplasmic inheritance for cv. ‘Dukat’ as maternal form and positive rij for the breeding clone no. 590 indicated the nuclear inheritance of morphogenetic ability. Cv. ‘Teresa’, as maternal component, showed nuclear inheritance of that trait in crossing with cv. ‘Dukat’ and with 590 cytoplasmic inheritance. The productivity of morphogenesis in strawberry depended on the parental combination and the direction of crossing.


INTRODUCTION
Most traits inherited by organisms are associated with genes located on chromosomes in cell nucleus.But sometimes the cytoplasm can infl uence the inheritance of some traits.That kind of heredity is called cytoplasmic inheritance (extrachromosomal inheritance, extranuclear inheritance) and it is caused by plasmogenes which are largely located in mitochondria or plastids.These two organelle genomes coexist in the cytoplasm of their cells: the mitochon-formation of whole plants, provided opportunities for numerous applications of in vitro plant biology in studies of basic botany, biochemistry, propagation, breeding, and development of transgenic crops (P h i l l i p s , 2004).
The aim of this experiment was to determine the infl uence of genes located in the cytoplasm on productivity of morphogenesis in Fragaria x ananassa Duch.species.

Plant material
The cultivated strawberry Fragaria x ananassa Duch.(2n=8x=56) belongs to the Rosaceae family.It was described and named in 1766 by the French botanist Duchesne.The species originated from chance hybridizations that occurred in some European botanical gardens between two octopliod species Fragaria virginiana and Fragaria chiloensis, imported from the New World at the end of the 16 th century and the middle of 18 th century, respectively.The genomic constitution of the parental species remains unknown (L e rc e t a u -K ö h l e r et.al. 2003).
The experiment was conducted in two stages: in the fi eld and in a plant tissue culture laboratory.The objects of the research were two cultivars of Fragaria x ananassa Duch.species: 'Teresa', 'Dukat' and the breeding clone no.590.The origin of these cultivars and clone is given in Tab. 1.

Obtaining the hybrid material of progeny F 1 from diallelic crosses according to Griffi ng's method 3
The hybrid material of progeny F 1 was obtained via Griffi ng's method 3. It relies on single and reciprocal crosses without progeny from self-pollination.The positive value of reciprocal cross effects (r ij ) shows chromosomal inheritance and the negative value -cytoplasmic inheritance of morphogenetic ability 1956).The diagram of these crosses is given in Tab. 2. In order to do those crosses, 60 fl ower buds on each maternal form were emasculated -30 for each paternal form.The emasculated maternal fl owers were pollinated by two paternal forms -30 maternal fl owers for each paternal form.Pollen was derived from newly opened fl owers and it was kept for about seven days in a desiccator at 0+4°C.The pollinated maternal fl owers were isolated properly and protected from environmental factors.The hybrid material of six progeny combinations F 1 was obtained from nearly ripe fruits.

Estimation of in vitro morphogenetic ability of F 1 generation
The morphogenetic ability of each progeny F 1 was estimated on the basis of the mean number of microplants, obtained from explants of progeny F 1 in two subsequent subcultures.
After the superfi cial sterilization in sodium hypochlorite, the seeds of the progeny combination F 1 were placed on MS medium (Murashige and Skoog, 1962) without growth regulators and with a limited concentration of sucrose and pH=5.7 (Tab.3).The achenes were sterilised for 2h, after that three to fi ve times rinsed in sterile distilled water under aseptic conditions.
Petri plates with seeds were located at temp.0+4°C for eight weeks in order to undergo stratifi cation.After stratifi cation, the plates were kept in a phytotron under controlled light conditions (16h day/ 8h night) and temp.18-20°C.The germination of the seeds was observed.The seedlings at the cotyledon stage or at the 1-2 leaf stage were placed on sterile MS medium with growth regulators in order to induce in vitro morphogenesis (the medium composition is given in Tab. 3).
Each progeny combination was represented by 40 seedling explants.Each seedling explant was numbered to control further proliferation in two subcultures.The mean number of microplants per explant was estimated for each of six progeny combinations.
Table 1 The origin of the analyzed breeding clone and strawberry cultivars.After eight weeks, the number of microplants derived from each seedling explant was counted.One microplant derived from each seedling explant was chosen randomly (40 from each parental combination) and placed again on sterile MS medium with growth regulators.After eight subsequent weeks, the number of microplants produced by individual explants was estimated again.In the experiment, 480 explants were proliferated.

Statistical analysis of obtained results
The results of the study were statistically evaluated by means of parameters used in estimating the inheritance of quantitative traits.The usefulness of parental forms for transmitting the morphogenetic ability to the progenies was estimated on the basis of general combining ability (GCA).

RESULTS AND DISCUSSION
Mean microplant number per explant between two subsequent subcultures varied in the analyzed progeny F 1 .There was observed an increase in the number of microplants during the second explant proliferation (Tab.4).In general, the total mean number of microplants per explant reached the value of 8.0 in the fi rst proliferation.In the second proliferation, a signifi cantly higher number of microplants per explant was observed (13.0).In the progeny F 1 of the parental combination 'Dukat' x 590, the number of microplants per explant was the lowest (5.0) in the analyzed plant material after the fi rst subculture.The highest number of microplants was produced by the explants of combination 590 x 'Teresa', both in the fi rst and second proliferation (Tab.4).Only in the F 1 progeny combination of the parents 'Dukat' x 'Teresa', the number of microplants produced per explant during the second subculture was almost two times lower (6.0 microplants) compared to the fi rst one, where 11.0 microplants per explant were observed (Tab.4).
The average values of the number of microplants after two subsequent subcultures for six proge-   nies F 1 and maternal/paternal groups were also significantly varied (Tab.5).The highest differences in mean number of microplants were observed in case of the analyzed progeny F 1 , obtained after crossing the breeding clone no.590 as a maternal form and 'Dukat' or 'Teresa' as paternal forms (11.0 and 17.0 microplants, respectively).In the reciprocal crosses between those two cultivars as maternal forms and the breeding clone no.590 as a paternal form, the number of produced microplants per explant was 7.0 and 11.0, respectively.The lowest insignifi cant differences in mean number of microplants were observed in the case of the explants obtained after the crossing of cv.'Dukat' and 'Teresa' as maternal forms with two paternal genotypes (Tab.5).On the other hand, the average values of the analyzed trait observed in the reciprocal crosses for cv.'Dukat' used as a maternal form in single crosses with the breeding clone no.590 and cv.'Teresa' were signifi cantly higher (11.0 and 10.0, respectively).Furthermore, in the case of cv.'Teresa' used as a maternal form in a single cross with the breeding clone no.590 in the progeny F 1 , the same tendency was observed.
The mean number of microplants estimated for the explants obtained in the progeny F 1 of the reciprocal crosses 'Teresa' x 'Dukat' was signifi cantly lower (10.0 and 8.0, respectively).The variability of the number of microplants obtained from the explants of the parental combinations 590 x 'Teresa' and 'Teresa' x 590, estimated by the coeffi cient of variability, reached the value below 50% (Tab.6).The highest coeffi cient of variability (over 60%) was estimated for 590 x ' Dukat' and 'Teresa' x 'Dukat' parental combinations (Tab. 6).That indicates that the parental combination and the direction of crossing must be taken into account.However, certain crosses may be better for one trait, but not necessarily as good for other traits (L i n and G e n g , 2008).
The positive effect of GCA was obtained in case of the breeding clone no.590 (1.496) and a high positive effect in case of cv.'Teresa' (1.746), whereas in cv.'Dukat' negative (-3.242).If a particular cultivar has a high GCA effect for a trait with a desirable high level, it means that the cultivar would be a valuable parental form in a breeding program designed to improve that trait (M a s n y et al. 2005) and the use of these parental forms should signifi cantly increase the value of the progeny (Ż u r a w i c z , 1990).But L i n and G e n g (2008) suggest that some parental lines with greater combining abilities in one trait may not be so good in another trait.
According to U b y s z -B o r u c k a et al. (1985), the negative effect of reciprocal crosses was used to indicate cytoplasmic inheritance.In case of cv.'Teresa' crossed with cv.'Dukat', the effect of reciprocal crosses was positive (Tab.7).On the other hand, the same maternal form crossed with the breeding clone no.590 revealed a negative value of r ij .The obtained results showed the negative effect of r ij for cv.'Dukat' used as a maternal form, both in the case of crossing with the breeding clone no.590 and cv.'Teresa' as paternal forms.The opposite situation was observed for 590 as a maternal form crossed with 'Dukat' and 'Teresa' as paternal forms, where the effects of reciprocal crosses showed a positive value (Tab.7).

CONCLUSIONS
1) The infl uence of plasmogenes on in vitro productivity of morphogenesis should be recognised in Fragaria x ananassa Duch.species.2) The morphogenetic ability of strawberry explants depended on the parental combination and the direction of crossing.
3) The effects of reciprocal crosses depended mainly on the choice of the maternal component.4) The best maternal component in the analyzed parental forms was the cultivar 'Teresa' which showed a high and positive value of general combining ability (GCA).5) The high and negative effect of GCA evaluated for cv.'Dukat' indicated this maternal form as the worst one.6) Unstable inheritance of morphogenetic ability was observed in cv.'Teresa'.7) The cytoplasmic inheritance of morphogenetic ability was assessed in cv.'Dukat' through the high and negative effect of reciprocal crosses.8) In the analyzed explants of the progeny F 1 , there was a tendency to increased productivity of morphogenesis in strawberry during the second subculture.
= Paula x (Senga Sengana x Talisman) Koralowa 100 x Gorella Redgauntlet S 1 x Senga Sengana S 1 The calculated effects of reciprocal crosses (r ij ) for single crosses i x j (i<j) were used to indicate those maternal forms which transmitted the morphogenetic ability extrachromosomally by plasmogenes (U b y s z -B o r uc k a et al. 1985; H a k i z i m a n a et.al. 2004; M a sn y et al. 2005).

▪
Means followed by the same letter are not signifi cantly different at P=0.

Table 2
Diagram of diallelic cross confi guration by Griffi ng's method 3 (single crossing x and reciprocal crossing xx without progeny from self-pollination).

Table 3
Composition of standard MS medium for stratifi cation of strawberry sowing material and strawberry micropropagation.Mean number of microplants per explant in the fi rst and second proliferation of the analyzed progeny F 1 .
▪ Means followed by the same letter are not signifi cantly different at P=0.05Table 5Average values ( ) of the analyzed trait after two subsequent subcultures for six progenies F 1 and maternal ( ♀• ) /paternal groups ( • ♂ ).

Table 6
Variability of the analyzed trait of six progenies F 1 .General combining ability (GCA) of parental forms and effects of reciprocal crosses (r ij ).