Response of selected genotypes of white and water yam to somatic embryogenesis

The shortest days to callus formation (15 d) were observed in Asiedu, which was significantly faster than the days taken by Swaswa by 5 d and TDa2014 by 11 d (Table 1). There were no differences between Kpamyo, Asiedu, and Ekiti2a in the percentage of callus formation with ≥ 95%. This is 47% higher than Swaswa and 62% higher than TDa2014. The number of plantlets regenerated was significantly higher than the rest of the genotypes. The plantlets were regenerated in Ekiti2a within a period of 58 d and this was significantly faster than the days from culture to regeneration in Asiedu and Swaswa by 8 and 10 d, respectively. The number of roots formed by Asiedu, Kpamyo, and Ekiti2a was not significantly different from each other, but they were statistically higher than Swaswa and TDa2014.

Table 1. Mean values of somatic embryogenesis traits in five genotypes of Dioscorea rotundata Poir. and Dioscorea alata L Relative multiplication rates of yam propagated via somatic embryogenesis and organogenesis

The average number of nodes obtained from the cultured explant at sixteen (16) wk of culturing via organogenesis ranged from 16 nodes (Asiedu) to 25 nodes (Ekiti2a and TDa2014); however, through SE, the average number of nodes ranged from 10 nodes (TDa2014) to 21 nodes (Asiedu) (Table 2). Between the organogenesis and SE propagation methods, the average number of nodes obtained in the five genotypes via organogenesis (21) was 24% higher than the average nodes obtained in the genotypes via SE (16).

Table 2. Propagation rates of five Dioscorea rotundata Poir. and Dioscorea alata L. genotypes regenerated through somatic embryogenesis and organogenesis Histology of callus tissues at 4, 6, and 8 wk of culturing

The photomicrograph of the three stages (4, 6, and 8 WAI) of somatic embryogenesis processes of the three white yam and two water yam genotypes revealed that at 4 WAI, there was no definite tissue arrangement in all the genotypes, rather an active cellular proliferation with no definite tissue arrangement pattern. However, at 6 and 8 WAI, there was structural tissue arrangement with visible vascular bundles (Fig. 1).

Figure 1.

Photomicrographs of somatic embryogenesis phases of white and water Dioscorea rotundata Poir. and Dioscorea alata L. at × 10. Keys: (a–c) Callus sections of Swaswa at 4, 6, and 8 wk of culturing respectively. (d–f) Callus sections of TDa2014 at 4, 6, and 8 wk of culturing respectively. (g–i) Callus sections of Asiedu at 4, 6, and 8 wk of culturing respectively. (j–l) Callus sections of Kpamyo at 4, 6, and 8 wk of culturing respectively. (m–o) Callus sections of Ekiti2a at 4, 6, and 8 wk of culturing respectively. Arrow: Vascular tissue arrangement.

Post-flask performance of somatic embryogenesis– and organogenesis-produced plantlets of white and water yam

Asiedu produced the highest number of new leaves (4.00 ± 2.34) during acclimatization, which was not significantly different from Ekiti2a (3.70 ± 2.15), Kpamyo (3.20 ± 1.94), and Swaswa (3.20 ± 1.23) but was significantly higher than TDa2014 (2.40 ± 1.47). The number of nodes produced by Asiedu (6.60 ± 3.69) was not significantly different from TDa2014 (6.55 ± 2.67), but they were both significantly higher than Kpamyo (4.45 ± 2.19). The height (10.02 ± 7.07) of Asiedu was significantly higher than the rest of the genotypes evaluated. The survival rates of the plantlets produced through somatic embryogenesis and organogenesis were not significantly different. However, the number of new leaves (3.96 ± 1.89), number of nodes (6.44 ± 2.67), and the height (8.73 ± 2.26 cm) of the plants produced through somatic embryogenesis were significantly higher than plantlets produced via organogenesis with 2.64 ± 1.71, 5.36 ± 2.83, and 5.33 ± 2.26 cm for number of new leaves, number of nodes, and the plant height, respectively. The interaction between genotypes and propagation pathways was significant for the number of leaves, number of nodes, and plant height except for the hardening survival rate (Table 3). The number of nodes produced after potting increased gradually from 4 to 8 wk in all the genotypes. At eight (8) WAP, Asiedu had the highest number of nodes (75.70 ± 35.16), which was significantly higher than the rest of the genotypes. The number of tubers (2.45 ± 1.39) produced by Asiedu and the average tuber weight (68.36 ± 20.79), respectively, were significantly higher than the rest of the genotypes. The number of tubers produced by TDa2014 (1.75 ± 0.72) was not significantly different from Kpamyo (1.55 ± 0.60) and Ekiti2a (1.30 ± 0.73) but was significantly (P ≤ 0.05) higher than Swaswa (1.15 ± 0.49), while the weight of tubers produced by Asiedu (68.36 ± 20.79) was significantly (P ≤ 0.05) higher than the other genotypes. However, the number of tubers produced by the plantlets raised through organogenesis (1.86 ± 1.11) was significantly higher than the somatic embryogenesis–raised plantlets (1.42 ± 0.70). The interaction between the genotypes and propagation pathways was significant (P ≤ 0.05) for the number of nodes and tuber weight (Table 4).

Table 3. Ex vitro performance of five (5) Dioscorea rotundata Poir. and Dioscorea alata L. genotypes propagated via somatic embryogenesis and organogenesis at 2 wk after acclimatizationTable 4. Relative post-flask growth performance and tuber yield of 5 Dioscorea rotundata Poir. and Dioscorea alata L. genotypes propagated via somatic embryogenesis and organogenesis at 4 to 8 wk after acclimatization and at harvest Variations in post-flask growth and yield performance among genotypes of white and water yam produced via different micropropagation pathways

The average number of new leaves, number of nodes, number of leaves, and plant height after 2 wk of acclimatization varied across genotypes and source of plantlets (organogenesis and SE) (Fig. 2). In Kpamyo, the number of new leaves produced by the plantlets regenerated through SE (3.8) was 32% more than the new leaves produced by Kpamyo plantlets regenerated through organogenesis (2.6). In Asiedu, the number of new leaves produced by the plantlets regenerated through SE (5.9) was 64% more than the new leaves produced by Kpamyo plantlets regenerated through organogenesis (2.1). New leaf production in Ekiti2a multiplied via SE (4.1) was 20% more than organogenesis-raised plantlets (3.3). In Swaswa, the new number of leaf production in the SE-raised plantlets (3.2) was 50% more than the organogenesis-raised plantlets (1.6). However, in TDa2014, the number of new leaves produced by the organogenesis-raised plantlets (3.6) was 22% more than the SE-raised plantlets (2.8). On the NON, in Kpamyo, Asiedu, and Swaswa, the nodal production in the SE-raised plantlets was 16%, 64%, and 14%, respectively, more than the organogenesis-raised plantlets. However, in Ekiti2a and TDa2014, the NON produced in the organogenesis-raised plantlets were 5% and 32% higher than the SE-raised plantlets, respectively. In the NOL produced by Kpamyo and TDa2014, the organogenesis-raised plantlets were 3% and 30%, respectively, higher than the SE-raised plantlets. On the plant height, the SE-raised plantlets were taller than the organogenesis-raised plantlets in Kpamyo, Asiedu, Ekiti2a, Swaswa, and TDa2014 by 14%, 69%, 1%, 43%, and 22%, respectively.

Figure 2.

(A) Number of new leaves, (B) number of nodes, (C) number of leaves, and (D) plant height of white and water Dioscorea rotundata Poir. and Dioscorea alata L. plantlets produced through somatic embryogenesis and organogenesis after 2 wk of acclimatization. Keys: NNL number of new leaves, NON number of nodes, NOL number of leaves, PH plant height, Org organogenesis, SE somatic embryogenesis.

There was gradual increase in the number of nodes produced by all genotypes in both organogenesis and somatic embryogenesis from the 2nd to 8th WAP (Fig. 3). At 8 wk of potting, the NON produced by SE-raised plantlets in Kpamyo, Ekiti2a, and Asiedu were 8%, 26%, and 28% higher than the organogenesis-raised plantlets, respectively. However, in TDa2014 and Swaswa, the NON produced by the organogenesis-raised plantlets were 4% and 13% higher than the SE-raised plantlets, respectively. There was 100% survival of the somatic embryogenesis–produced plantlets after hardening in all genotypes while in the organogenesis-raised plantlets, all the plantlets introduced to hardening survived except for Asiedu and Kpamyo that had 90% and 80% survivals, respectively (Fig. 4A). The number of tubers by the organogenesis-raised plantlets after 7 mo of potting in Kpamyo, Ekiti2a, Asiedu, TDa2014, and Swaswa was 6%, 27%, 42%, 16%, and 8% higher than the SE-raised plantlets, respectively. The number of tubers by the organogenesis-raised plantlets after 7 mo of potting in Kpamyo, Ekiti2a, Asiedu, TDa2014, and Swaswa was 6%, 27%, 42%, 16%, and 8% higher than the SE-raised plantlets, respectively. In addition, the weight of tubers harvested from the organogenesis-raised plantlets after 7 mo of potting in Ekiti2a, Asiedu, TDa2014, and Swaswa was heavier than the SE-raised plantlets by 52%, 4%, 54%, and 11%, respectively. However, in Kpamyo, the weight of tuber produced by the SE-raised plantlets was 21% heavier than the organogenesis-raised plantlets (Fig. 4B). The sequence of activities (stages of regeneration) in the SE system started from the induction of embryogenic callus from axillary bud explant, followed by the production of somatic embryos and the maturation and production of plantlets from the somatic embryos (Fig. 5A–C). The plantlets obtained via SE and organogenesis were successfully acclimatized and potted. This led to the production of minitubers at 7 mo after potting from the plantlets obtained from both propagation pathways (Figs. 4 and 5).

Figure 3.

Average number of nodes produced by somatic embryogenesis– and organogenesis-raised plantlets of white and water Dioscorea rotundata Poir. and Dioscorea alata L. in post-flask. Keys: NON2–NON8 number of nodes at 2, 4, 6, and 8 wk after potting, Org organogenesis, SE somatic embryogenesis.

Figure 4.

(A, B) Post-flask survival and tuber yield of in vitro produced plantlets of white and water Dioscorea rotundata Poir. and Dioscorea alata L. Keys: NoIntro number of plantlets introduced to hardening, NSH number of plantlets successfully hardened, NOT number of tubers harvested after 7 mo, Org organogenesis, SE somatic embryogenesis.

Figure 5.

Stages of seed Dioscorea rotundata Poir. and Dioscorea alata L. tuber production via somatic embryogenesis in Asiedu. (A) Callus induction from the axillary bud (AB) explant at 2 wk of culturing (WOC). (B) Callus cultures at 4 WOC. (C) Somatic embryos at 6 WOC. (D) Regenerated plantlets at 10 WOC. (E) Regenerated plantlets at 17 WOC. (F) Hardening of plantlets at 17 WOC. (G) Hardened plantlets at 19 WOC. (H) Potted in vitro plantlets at 27 WOC. (I) Harvested tubers at 45 WOC. Arrow: Torpedo-shaped somatic embryo.