Abstract:
Cactus pear [(Opuntia ficus-indica (L.)MILL] is a horticultural crop which is extensively used as food for humans and feed for animals in many countries of the world. The plant has widely adapted to different parts of Ethiopia and is an integral part of the culture and economy of Tigray, Northern Ethiopia. Although, Cactus pear can be sexually and asexually propagated, propagation has its own limitation such as genetic segregation, a long juvenile stage, and slow growth, material require large spaces for propagation. Therefore, to overcome these propagation barriers and to fulfill the demand for large-scale cultivation in a short period by rapid mass multiplication, experiments were conducted with the aim of developing a protocol for the in vitro propagation of cactus pear from in vitro derived areole explants and selection of cactus pear for salinity and drought tolerance at the Plant Tissue Culture Laboratory of Mekelle Agricultural Research Center, Northern Ethiopia. The experiments were laid out in completely randomized design (CRD) with three replications. For the first and second experiment, simple and reliable strategy was followed for micro-propagation of spineless and spiny cactus pear in the presence of the single growth regulator, Kinetin (kin), at four different concentrations (1, 2, 4 and 8 mg/l) and at three different sucrose concentration (20, 30, 50 g/l) was investigated for induction of root. Results of the first experiment revealed that the highest values for number of shoots for all genotypes and length of shoot was recorded on medium supplemented with 2 mg/l kin and length Kin, respectively. The second experiment, the highest values (P ≤ 0.05) for number of shoots and shoot length for all genotypes recorded on MS medium supplemented with 1mg/l Kin, For both experiments ( 1 and 2) the shoots (4-5 cm length) were cultured on half-strength medium supplemented with sucrose concentration of 30 g/l exhibited the maximum root induction for all genotypes. Regarding root length, root fresh weight and dry weight, MS medium supplemented with 50 g/l of sucrose had obtained the highest value. For acclimatization, culturing the in vitro seedlings on sand, cocopeat, and compost or a mixture of the three at the ratio of 2:1:1 gave the highest survival capacity (100%). In the third experiment, six genotypes namely, Suluhna, Gerao, Lemats Beles, Keyih Beles, Limo and Shenkor were used to study the performance of cactus genotypes for salt tolerance. Callus was initiated in MS medium at 4 mg/l Dichlorophenyl Acetic Acid ( 2,4- D ) + 0.5 mg/1 6- Benzyl Adenine (BA) and at different concentration of Sodium chloride (NaCl) (0 mM, 50 mM, 100 mM, 150 mM and 200 mM ) were added to the medium to create salt stress .Among the xviii six genotypes plated in MS medium with 0 mM NaCl or supplemented with 50 mM NaCl, Suluhna was significantly (p ≤ 0.05)superior for callus induction (100%), highest callus fresh weight (4.91g). The highest level of shoot regeneration was observed in Suluhna (41.6%) followed by Gerao and Keyih Beles (33.3%). A decreasing pattern in number of shoot of the cactus genotypes were observed with increasing salinity levels and adversely affected at 100 mM NaCl. The root fresh weight, root dry weight root number and length decreased significantly from (P ≤ 0.05) 50 mM NaCl and no rooting occurred when explants were grown with 100 mM NaCl. In the fourth experiment, the non-ionic water soluble polymer polyethylene glycol (PEG) of molecular weight 6000 was used as osmoticum to simulate water stress at 0, 10, 20 and 40 g/). In the first culture, the MS medium was supplemented with (2, 4-D (4 mg/l) and BA (0.5 mg/l for callus induction and was semi solidified with 0.8% agar and 30g of sucrose. Significant differences were observed among the genotypes, treatments and their interactions for the evaluated growth parameters. At 10 g /l PEG, the callus induction frequency , callus fresh weight and plantlet regeneration were highest (83.3 %, 5.5g and 66.7%, respectively) in genotypes Suluhna followed by Gearo. At 40 g/l PEG, callus induction frequency, callus fresh weight and plantlet regeneration were highest in Suluhna (41.7%, 2.75g and 41.7%) respectively. However, Shenkor and Keyhi Beles was induced callus that became reddish black within 35 days in medium with 40 g/l PEG. Both shoot and root production decreased with increased PEG level in the medium. At 40 g/l PEG in MS medium, the highest shoot number was in Suluhna genotype (4.33) followed by Gearo (3.67). The highest shoot length was in Suluhna (2.11cm) with no significant difference with Gerao (2.02cm) while root number (5.00) and root length (1.41cm) were in Suluhna. In conclusion, it is beneficial to use the in vitro propagation and selection for drought and salinity protocols developed in this study for mass micropropagation and overcome the challenges of conventional propagation of cactus pear.