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The effect of the interaction of genotype and culture medium on the initiation of callus from immature embryos and subsequent plant regeneration was investigated in eight hexaploid wheat lines. Intervarietal differences in culture response and interaction of the genotype with coconut milk are reported. The relative contributions of media and genotype effects to culture performance are assessed. The observation that primordia and shoot development was promoted by coconut milk in some lines and inhibited in others is particularly significant given that coconut milk is widely used to try to improve culture response. This report shows that this effect is dependent on the genotype of the tissues in culture.

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A technique for the in vitro colchicine treatment of microspore-derived haploids of rape {Brassica napus L.) is presented. This allows effective and timesaving chromosome doubling. The most effective method consists of an application of 50 mg/1 colchicine in a liquid B5 medium to the rooted microspore-derived plantlets for 4-8 days. By this treatment more than 50 % of the plants developed at least diploid sectors. A change of medium after some days may even increase this diploidization rate.Since the basic investigation of LICHTER (1982) microspore culture became the most common technique for the production of haploids in rape {Brassica napus L.). Even large scale embryoid production and plantlet regeneration are now possible (POLSONI et al. 1988). For breeding purposes and progeny analyses it is necessary to double the chromosome number of the haploid plantlets. Only a small number, about 20-30 % of the regenerates, was shown to undergo spontaneous doubling (LICHTER et al. 1988). So far chromosome doubling is usually carried out by treatment of the plantlets after transfer to soil, either by injecting a 0.2 % solution of colchicine into the secondary buds (LICHTER et al. 1988) or by submerging the roots into a 0.1-0.2 % solution of colchicine for 5-6 hours (POLSONI et al. 1988, SwANSON et al. 1989). Both techniques require previous transplantation of the plantlets into soil and are, therefore, time-consuming. The present study suggests an in vitro application of colchicine which is quite effective and allows the production of diploids in less time.For the production of the haploid material, donor plants of a microspore-derived homozygous diploid line originating from the spring rape variety 'Duplo', were grown in the greenhouse in early 1989. Microspore culture and plantlet regeneration were executed as previously described (MATHIAS 1988). At the 3-4 leaf stage groups of 1-3 rooted plantiets were aseptically transferred to sterile 50 ml culture flasks. The roots of the plantlets were immersed in 5-10 ml liquid B5 medium (GAMBORG et al. 1968) with an addition of 2 % sucrose and 10-100 mg/1 colchicine ( Table 1). The basic medium was autoclaved and then 1-10 ml/1 of a sterile, filtered 1 % (w/v) stock solution of colchicine were added. After 1-8 days of stationary culture at 25 °C in a 16 hrs photoperiod the colchicine medium was removed and replaced by 5-10 ml of the basic B5 medium. After an additional culture of 1-2 weeks under the same conditions as above, plantlets were transplanted to soil (see MATHIAS 1988). The ploidy was determined by evaluating morphological characters of the flowers, e.g. size and pollen production. U.S. Copyright Clearance Center Code Statement: 01 79-9541/91/0601-0082$02.50/0

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