In vitro Morphogenesis and Microtuber Induction in Potato (Solanum tuberosum L.)

Abstract

Establishment of an efficient tissue culture system is an important step for plant breeding programs. Initially tissue culture protocols have been established for four commercially important potato cultivars namely, Diamant, Almera, Alpha, and Agria. Seed tubers were incubated at 4'C under dark condition for sprouting and used for in vivo culture. Bud, node, stem segment, leaflet segment, and tuber segment explants were sterilized using sodium hypochlorite. Nodal explants were used for establishing sterile in vitro micoplants, rapid multiplication, and subsequent rooting, acclimatization, microtuber induction and indirect plantlets regeneration. For multiplication response for all cultivars and all hormones treatment regeneration percentage was ranging between 25% - 100%. For shoot number, Almera gave the maximum number of shoot (5.4:l:O.7lI shoot/explants) in MS media supplemented with 3.0 mg/l TDZ+0.1 mg/l NAA. For rhizogenesis of direct regenerated micrpoplants, it was observed that when MS media without auxin was used (control) 100, 90%, 50% and 50% Of the shoot produced root with root number average 4.0iO.9, 2.11 0.6, 1.0=t0.4 and 1.0i0.4 roots/shoot and root length 8.8il.3, 2.8=l=O.8, 3.7:!: 1.5 and 3.5=hl.4 from cultivar Almera, Diamant, Alpha and Agria respectively. When MS media supplemented with various auxin (NAA, IAA, and IBA) was used, it was observed that 100% root formation from all cultivars with high root number per shoot ( as compare to control) was obtained with maximum root number of 35.0i1.3 roots/ shoot by using IBA at 1.0 mg/1 for cultivar Almera, 19.9i2.4 roots/ shoot by using IBA at 0.5mg/l for cultivar Diamant, 5.5:+: 0.9 roots/ shoot by using (IBA2.0 or IAAl.5) for cultivar Alpha, and 5.0:*:l.5a roots/shoot by using IBA 2.0mg/l for cultivar Agria. The best acclimatization response (95%) was obtained in sterilized soil, 50% clay: 50%. Microtuberizations was carried with Diamant and Almera. Highest number of microtubers/jar (6.0:l:0.5microtubers/jar) was yielded by Almera in MS media fortified with sucrose 8% without hormone, at dark On the other hand maximum microtuber weight, was given by Almera cultivar in MS media supported with 60 g/l sucrose without hormone which is (1250.3il3.0 mg/tuber) in dark. For callus induction, the effect of explants explained that, among all explants the tuber segment was found to be most effective for callusing degree in all cultivars. , highest degree of callus (6.0:k0.0) was induced by Diamant on 2, 4-D at 3.0 mg/l, To evaluate the effect of growth regulators on callus induction from tuber segments explants, different of growth regulators were used. When MS media supplemented with IAA, IBA, and Kin used there was no callus induced in all cultivars. Highest callus degree (6:t0.0) was induced for Diamant on media supplemented with 3.0 mg/1 2, 4-D and2.0 mg/l 2, 4-D in combination with BAP at 2.0 mg/l. For cultivar Agria the higher callus degree recorded (6.0:k0.0a) was obtained on MS media supplemented with 2.0 mg/l 2, 4-D in combination with (either 2.0 mg/l or 5 mg/1 BAP) or BAP at 5.0 mg/l in combination with 2, 4-D at (1.5mg/1 or 3 mg/1). For Almera higher degree of callus (6.0i0.ll and 5.8i0.l“) was obtained by using 2.0 mg/l of BAP in combination with 2,4-D at three levels (l.0,l.5 and 2 mg/1 2,4-D ) , 2,4-D at 2.0 mg/l in combination with BAP at 2.0 and5.0 mg/l and 2,4- D at (1.5 and 1.0 mg/l), in combination with BAP at 5.0 mg/l. Higher degree of callus (5.3=t:0.2”)for Alpha was obtained on MS media supplemented with 1.5mg/l and 2.0mg/l 2,4-D in combination with 5.0mg/l and 2.0 mgl BAP respectively. Callus was induced with all types of media with variation among cultivars, it wasl/2 B5 and MB with degrees of callus (4.6i0.2 and 4.3:k0.2) respectively for Diamant. For Agria higher callus degree was induced by l/2 B5 with degree of callus (4.0:k0.2). Lowest callus degree was obtained by Alpha there was no significantly different between media for callus induction with all using media. The Almera cultivar induced same callus degree with 1/2MS, l/2 B5 and B5 treatments. Regeneration of indirect microplant, explained that TDZ and BAP alone and in combination with auxin mainly (IBA) produced shoot with wide variation between the three cultivars, TDZ 5.0mg/1 and BAP 4.0 mg/l in the cultivars Diama.nt induced highest number of shoot (3.4=t:0.9 and 2.9i0.6“ shoot/callus). The highest root number was achieved by Alpha (42.5=t:3.4 root/ plant) with 100% rooting in half MS media fortified with 2.0 mg/l IBA and (38.8=t:l.6 root/plant) obtained in half MS media supplemented 1.5 mg/1 IBA. For the same cultivar maximum root length was obtained by half MS media without growth regulators, with average length (8.3:l:l.0 cm). For acclimatization of indirect regeneration rooted plantlets, 95% survival rate was obtained when sterilized soil, clay: sand (1: 1 v/v) was used tmder greenhouse conditions.