Micropropagation of thirteen Malus cultivars and rootstocks, and effect of antibiotics on proliferation

Abstract

Several factors that affect in vitro establishment, proliferation, and rooting of thirteen Malus cultivars and rootstocks were studied . Apple shoot tips (1 .5 ± 0.5 cm in length) were established using ascorbic and citric acids as antioxidants . Four proliferation media containing 1 .0 mg 1 -1 BA and different concentrations of IBA and GA3 were tested. Proliferation rates varied depending on the genotype and medium used . The highest proliferation rate was obtained for a rootstock that produced 11 .6 ± 2 .5 shoots (1 .5 ± 0 .8 cm in length) per tube per month . Rooting was induced with IBA for all the genotypes tested . The optimal IBA concentration was cultivar dependent (between 0 .1 and 1 .0 mg 1-1 IBA), and lower concentrations were necessary to induce rooting in liquid rather than in solid medium .The effects on shoot-tip proliferation of cefotaxime, carbenicillin and kanamycin, three antibiotics commonly used for transformation studies, were also evaluated . Cefotaxime at 200 mg 1 -1 stimulated shoot growth and development, but at 500 mg 1 -1 caused abnormal shoot morphology. Carbenicillin at 500 mg 1 -1 , alone or in combination with cefotaxime at 200 mg 1 -1 , inhibited proliferation and caused excessive enlargement of the basal leaves, inducing callus formation and release of phenolic compounds in the medium . Kanamycin at 50 mg 1 -1 was phytotoxic and caused shoot chlorosis and necrosis . Consideration of the toxicity of these antibiotics is critical when designing transformation schemes for selection and recovery of transgenic apple plants .Abbreviations: BA = benzyladenine ; cef = cefotaxime ; crb = carbenicillin ; GA3 = gibberellic acid ; IBA = indole-3-butyric acid ; Kan = kanamycin ; ms = Murashige and Skoog [19] macro-and micro-nutrients ; NAA = naphthalene-acetic acid . 55

In vitro shoot multiplication as influenced by repeated subculturing of shoots of contemporary fruit rootstocks

“…4). According to Yepes and Adwinckle (1994), lack of vascular connections between roots and shoots was implicated in low survival of in vitro rooted apple plantlets after transfer to the soil. Similarly, indirect in vitro rhizogenesis through callus formation can be one of the reasons for low percentage of acclimatization in Gisela 5, as was shown in pear cultivar Bartlett (Bommineni et al 2001).…”

<i>In Vitro</i> Propagation and Recovery of Eight Apple and Two Pear Cultivars Held in a Germplasm Bank

“…[31]. The combined effect of BA and GA 3 for establishment of the in vitro culture of fruit trees has previously been reported for pear trees [14] and apple trees [21], even when starting from buds and apple tree nodes [33].…”

Dormancy development during cold hardening of in vitro cultured Malus domestica Borkh. plants in relation to their frost resistance and cryotolerance

“…Greensleeves were used. In vitro cultures were subcultivated on medium Murashige and Skoog (1962), which was slightly modified by authors Yepes and Aldwinckle (1994) and Paul T. Lynch (personal communication). Sorbitol (30 g l -1 ) was used instead of the normally used sucrose as carbohydrate source and phytohormones BAP (1 mg l -1 ), IBA (1 mg l -1 ) and GA 3 (1 mg l -1 ) were added to the media at a temperature of 23 ± 0.5°C, photoperiod of 16/8 (day/night) and light intensity of 120 lE s -1 m -2 PAR.…”