II. 3. Propagation of barley by in vitro culturing of lateral primordia of shoots and of fragments of rosette shoots.
M. D. Gaj, M. Maluszynski, M. Gaj, Department of Genetics, Silesian University, ul. Jagiellonska 28, 40-032 Katowice, Poland.
Efficiency of vegetative propagation of barley by callus cultures of somatic tissues of this plant is still very poor (Dale and Deambridgio, 1979; Orton, 1979). For this reason, an attempt has been undertaken to obtain barley plants by way of stimulating growth under in vitro conditions of primordia of lateral shoots as well as of fragments of rosette shoots.
Experimental material consisted of spring barley Hordeum vulgare L. var. distichon, Georgia variety and the intensive tillering 47 400 mutant, derived from the Delisa variety by treatment with MNUA. Explants (2-5 mm long primordia of lateral shoots and longitudinal fragments of the rosette shoot) were obtained from plants in their three-leaf stage, growing under sterile conditions on perlite with Linsmaier and Skoog nutrient medium (1965) containing micro-and macro-elements, half by half. Growth of explants was performed on media consisting of the micro- and macro-elements and vitamins of Norstog and Smith (1963), 8g 1-1 Difcobacto agar and a varying composition of sucrose and hormones. Medium SJ1 was supplemented by 30 gl-1 sucrose, medium SJ2 by 60 gl-1 sucrose and 1 mgl-1 GA3 and medium SJ3 by 30 gl-1 sucrose, l mgl-1 IAA and 10 mgl-1 kinetin. Each medium was adjusted to pH 5,6-5,8 with 1N NaOH and/or HC1. Cultures of the explants were maintained at 20-25°C, under 16-hour illumination per day and 80% relative humidity.
Growth of primordia of the lateral shoots and formation of green, rooted plants were gained after 8 weeks of culturing. The barley plants were then removed and placed on perlite in test tubes containing Linsmaier and Skoog medium (1965); they were replanted into pots filled with soil. In these pots the plants spread out intensely, made rapid growth, became visibly lush and set seeds as usual. A cytogenic analysis of PMC's stained by the Feulgen's reagent technique has unravelled a regular process of meiosis and a diploid number of chromosomes, 2n=14. In addition to the green, rooted shoots, in part of the explants there took place incomplete organogenesis consisting of formation either of a shoot (green or white) alone, or only of roots.
Efficiency of the method used for progagation of barley, its dependence on the composition of the nutrient medium and the type of the explant, is illustrated in Table 1. The findings indicate that a highest frequency of obtaining plants is to be gained on SJ1 and SJ2 media, by culturing primordia of lateral shoots. Very little success was observed on the other hand, on the same media, by culturing of fragments of rosette shoots. Moreover, it has been ascertained that success with the suggested method for propagating barley largely depends on the genotype of the plant from which the explants had been derived. In a total of 20 plants only 2 owed their origin to Georgia variety, while all the remaining ones were derived by culturing explants of mutant 47 400.
Table 2. Efficiency of propagation of barley by shoot tip culture.
With the purpose of increasing the number of primordia of lateral shoots obtained per plant, culture of shoot apex isolated from young seedlings was conducted according to the suggestion of Raman et al. (1980). The results obtained by this method are presented in Table 2.
It seems that efficiency of this suggested method will be liable to improvement by further elaboration of the conditions for cultivating the maternal plants.
References:
Dale, P.J. and E. Deambridgio. 1979. A comparison of callus induction and plant regeneration from different explants of Hordeum vulgare. Z. Pflanzenphysiol. 94:65-67.
Linsmaier, E.M. and F. Skoog. 1965. Organic growth factor requirements of tobacco tissue cultures. Physiol. Plant. 18:100-128.
Norstog, K. and J.E. Smith. 1963. Culture of small barley embryos on defined media. Science 142:1655-1656.
Orton, T.J. 1979. A quantitative analysis of growth and regeneration from tissue cultures of Hordeum vulgare L., H. Jubatum and their interspecific hybrid. Environmental and Experimental Bot. 19:319-335.
Raman, K., D.B. Walden and R.I. Greyson. 1980. Propagation of Zea mays by shoot tip culture: A feasibility study. Ann. Bot. 45:183-189.
