Micropropagation In Vitro Regeneration And Molecular Diversity Study Of Moringa Stenopetala (bakf .) From Ethiopia

The tree species Moringa stenopetala belongs to the family Moringaceae and representedrnonly by a single genus Moringa. M. stenopetala is a native tree in Southern Ethiopia,rnNorth Kenya and Eastern Somalia. It plays a vital role for household food security, asrnsource of income, medicine , purification of water, windbreak, fodder, fuel and shade treernall year round. Conventional propagation o/ M. stenopetala involves the use of seeds andrncuttings . However, the application of fast and efficient biotechnological methods tornpropagate and improve different traits of interest is much needed. The objectives of thisrnstudy were to develop micropropagation and in vitro regeneration protocols for Moringarnstenopetala and to analyze its genetic diversity using ISSR molecular marker. Shoot tipsrnobtained from in vitro germinated seedlings were used for culture initiation. Allrntreatments with BAP in combination with NAA showed 100% shoot initiation beginningrnfrom the first culture. BAP at lower concentration was found to be the best for shootrnmultiplication. Of the treatments with BAP, KN, TDZ, IBA and NAA used for shootrnmultiplication, high quality and maximum mean number (11.16+0.14 and 10.86^0.14 ) ofrnshoot per shoot tip explant were attained on medium containing 0.5 mg/l BAP and 1.0rnmg/I BAP with no significant difference. Similarly, application of 1.0 mg/l BAP resultedrnin the highest number of induced axillary shoots (13.66±0.20) per nodal explant. Halfrnstrength MS medium containing NAA combined with IBA was virtually better in promoting root induction than 1/2 MS medium containing IBA or NAA alone. The leavesrnof in vitro multiplied shoots were used to develop the in vitro regeneration protocol basedrnon using TDZ, 2,4-D and NAA alone or in combination. The use ofTDZ alone at lowerrnconcentrations and application of 2,4-D, TDZ and/or NAA together led to very strongrncallus formation on MS medium. The maximum shoot regeneration percentage (63.66%)rnwas achieved on a medium supplemented with BAP and KN each at 1.0 mg/lrnconcentration. A genetic diversity study based on ISSR was also conducted on a total ofrn150 samples representing 14 populations from different parts of Ethiopia. The highestrnlevel ofpercent polymorphism (53.66%) was obtained from samples of Arba Minch Betornarea and the least level of polymorphism (46.34%) was obtained from individuals in ArbarnMinch Merab Abaya area. The AMOVA results revealed that the genetic variation atrnpopulation level accounted 21.47%) of the total variation, while the within populationsrncomponent accounted for 78.53% showing a higher within population variation thanrnamong populations. Jaccard’s similarity coefficient for populations ranged from 0.601 torn0.749, with the highest similarity value (0.749) was shown between populations fromrnKonso town and populations from outlying areas of Konso. Findings from this studyrncould be used for mass propagation of disease free planting materials within relativelyrnshort period and contribute to further improvement of the plant. In addition, the resultsrnobtained here can contribute to food security of both human and cattle, solve the demandrnof plantlets both in quality and quantity and help to increase source of income for localrnresidents as well as contribute to the genetic improvement of the species.rnKey words: In vitro regeneration, inter simple sequence repeat (ISSR),rnmicropropagation, Moringa, node, rooting

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