Plant Cell, Tissue and Organ Culture 67: 267–270, 2001. © 2001 Kluwer Academic Publishers. Printed in the Netherlands. 267 In vitro zygotic embryo culture of wild Musa acuminata ssp. malaccensis and factors affecting germination and seedling growth MJ Asif, C Mak & RY Othman Institute of Biological Sciences, Division of Genetics and Molecular Biology, University of Malaya, 50603 Kuala Lumpur, Malaysia Received 1 August 2000; accepted in revised form 30 May 2001 Key words: banana, embryo culture, Musa Abstract In vitro zygotic embryo culture of wild banana significantly increased the germination compared to greenhouse grown seeds. Embryo orientation and BAP concentration significantly affected germination rate. These factors together with gelling agent, dark and light conditions and coconut water, also showed variable effects on the number of roots per plant, root length, shoot length, number of days to root emergence and number of days to shoot emergence. Introduction Banana and plantain cultivars are attacked by several pathogens throughout the world tropics. By far the most serious pathogen is Fusarium oxysporum f. sp. cubense. There is no known chemical or cultural con- trol for the pathogen. Hence, varieties attacked by this pathogen must be replaced by resistant varieties (Rowe, 1990). Malaysia is a center of diversity for both cultivated and wild bananas. The wild progen- itors of the cultivated bananas co-evolved with the Fusarium oxysporum f. sp. cubense (Vakili, 1965) of- fering a possible source of resistance to the disease. The wild progenitors of the edible bananas produce seeds while most of the edible clones are seedless with a few exceptions such as ‘Pisang awak’ subgroup ABB (Simmonds, 1966). Intractable fertilization barriers such as moderate to high levels of female sterility and triploidy make genetic improvement of parthenocarpic Musa clones slow and technically difficult. Hybrid plant production in the most common triploid clones is further complicated by low seed set and germination (Shepherd et al., 1987). This is due to seed mor- tality caused by endosperm failure. Embryo culture before embryo abortion could rescue these interesting crosses. Little is known about the factors that affect seed germination in Musa, except that germination is ex- tremely variable and relatively difficult to obtain un- der natural conditions. The use of banana seedlings as research tools and the increased emphasis on ba- nana breeding programs requires improved germin- ation rates. In the current studies, seeds of Musa acuminata subspecies malaccensis have been used to optimize in vitro culture of zygotic embryos and to identify factors affecting germination and seedling growth. Materials and methods Mature seed samples of a diploid Musa acuminata ssp. malaccensis produced by open pollination were used. The fruits were thoroughly washed with distilled wa- ter and then soaked in 1.4% (v/v) sodium hypochlorite solution for 30 min. The fruit skins were removed, and seeds were removed under axenic conditions. Seeds were planted in soil mixture (sand + leaf compost) and maintained in the greenhouse. For in vitro culture, seeds were soaked in 1.4% (v/v) sodium hypochlorite solution for 10 min fol- lowed by a quick wash with 70% (v/v) ethanol.