Table 1.
Factors involved and their influence on micropropagation of Adhatoda vasica (arranged in chronological order).
| Explant | Surface sterilization | Culture medium composition | Culture condition | Regeneration response | Acclimatization | Reference |
|---|---|---|---|---|---|---|
| Leaf (in vitro) | 0.1% HgCl2 for 5 min → 4 interim rinse with sterile water | MS + 1.0 mg/1 BA + 0.1 mg/1 NAA | 25 ± 1 °C temp, 60% RH, 16 h photoperiod, with an irradiance of 3000 lux | Maximum number of shoots with optimum callus growth | Regenerated plantlets were acclimatized in soil | [49] |
| PGR-free MS | Rooting | |||||
| Nodal segment | 1% Savlon 10 min → 80% ethanol for 30 sec → 0.1% HgCl2 for 7–10 min → 4–5 interim rinse with sterile water | MS + 0.5 mg/l BA + 0.1 mg/l NAA | 26 ± 1 °C temp, 60% RH, 16 h photoperiod, with an irradiance of 2000–3000 lux | Maximum 10.5 shoots of 4 cm and 2.8 of leaves/ shoot | 80% survival on garden soil, sand and compost (2:1:1) | [32] |
| PGR-free MS | 100% rooting, 3.5 roots/shoot with 4 cm length in 15 days | |||||
| Leaf | Not mentioned | B5 + 1 mg/l 2,4-D | Not mentioned | Callus induction | Wasn’t carried out | [41] |
| B5 + 0.1 mg/l Kn | Shoot regeneration | |||||
| Shoot tip | 2–3 drops teepol for 5–10 min → 0.01% HgCl2 for 7–18 min → thorough rinse with sterile water | MS + 0.5 mg/l BA + 15% CM | 25 ± 1 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 3000 lux | 4.3 shoots per explant with 5 leaves 3.2 nodes/shoot | 85% plants were acclimatized in sterilized sand and soil mixture (3:1) | [47] |
| PGR-free MS | 9.33 roots per shoot with 0.6 cm length | |||||
| Leaf | 5 drops Tween 80 for 5 min → 0.1% HgCl2 for 5 min → 3 rinse with sterile water | MS + 21.5 μM NAA + 19.7 μM IBA + 9.3 μM Kn | 25 ± 2 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 30 μE/m2/s PPFD | 76% callus with precocious roots | Wasn’t carried out | [42] |
| Petiole | MS + 4.5 μM 2,4-D + 2.3 μM Kn | 62% callus with precocious somatic embryos | ||||
| Nodal segment | 0.1% HgCl2 for 5 min → 4–5 rinse with sterile water; or, 0.1% HgCl2 for 5 min → Geneticin treatment → 4–5 interim rinse with sterile water | MS + 10 mg/l BA | 25 ± 2 °C temp, 80% RH, 16 h photoperiod, with an irradiance of 100 μmol/m2/s PPFD | 7.75 shoots/explant in 4 weeks | Successfully acclimatized for 3 weeks in soilrite with liquid ½ MS nutrient spraying | [8] |
| MS + 1 mg/l BA + 1 mg/l Kn | 30 shoots/explant in 6 weeks | |||||
| MS + 0.1 mg/l IBA | 90% rooting | |||||
| Shoot tip, Nodal segment | 1% Savlon (w/v) and 2 drops Tween 80 for 20 min → unspecified HgCl2 for 5 min | MS + 2 mg/l BA + 0.2 mg/l NAA | 25 ± 2 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 3000 lux | 90% of explants produced 7 shoots/explant with 4.9 cm length in 28 days | 80% plantlets were acclimatized in garden soil + cow dung + sand (1:1:1) | [35] |
| MS + 1 mg/l IBA | 80% rooting with 3–4 roots/shoot of 3 cm length were recorded in 28 days | |||||
| Leaf, Petiole, Nodal segment | Unspecified | MS + 10.7 µM NAA + 2.2 µM BA | Unspecified | 90% repeatability to induce callus with 7 day callus induction | Wasn’t carried out | [53] |
| Leaf | 0.1% HgCl2 for 5 min → 3 rinse with sterile water | MS + 1.5 ppm 2,4-D + 1.5 ppm IAA + 1.5 ppm Kn + 1.5 ppm BA | 24 ± 2 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 3000 lux | 75% callus induction and proliferation with 18.16 g fresh weight | Wasn’t carried out | [43] |
| Nodal segment, Shoot tip, Petioles, Leaf disc | 0.1% HgCl2 for 5 min → thorough rinse with sterile water | MS + 2.0 mg/l BA + 0.5 mg/l NAA | Unspecified | Maximum callus induction from nodal segments | Rooted plantlets were acclimatized in a mixture of sandy soil and FYM (1:1) | [38] |
| MS + 2.0 mg/l 2,4-D, 0.5 mg/l Kn and 0.5 mg/l GA3 | Embryogenic callus proliferation | |||||
| ½ MS + 0.5 mg/l IBA + 0.2 g/l AC | Rooting | |||||
| Shoot tip | Unspecified | MS + 22.20 µm BA | Unspecified | High frequency and maximum number of multiple shoots | The rooted plantlets were hardened and established at 50–60% | [50] |
| PGR-free MS | High frequency of rooting | |||||
| Axillary meristems, Leaf, Nodal segment | 0.1% HgCl2 for 2 min → thorough rinse with sterile water | MS + 1 mg/l BA + 1 mg/l GA3 | 25 ± 2 °C temp, unspecified RH, 10 h photoperiod, with unspecified irradiance | High frequency shoot multiplication | Wasn’t carried out | [36] |
| MS + 1 mg/l Kn + 2 mg/l 2,4-D 1 mg/l BA + 0.1 mg/l Pic | Callus initiation within 5 days, both friable and green calli | |||||
| Nodal segment | 0.2% HgCl2 for unspecified duration → 4–5 interim rinse with sterile water | MS + 15% (v/v) CW + 5 mg/l BA | 25 ± 2 °C temp, unspecified RH, 16 h photoperiod, with unspecified irradiance | 14 shoots with 3 cm long in 8 weeks | 80% plants were acclimatized under laboratory conditions → transferred to pots filled with sterilized soil: sand mixture (1:3) | [33] |
| MS + 1 mg/l IBA | Rooting (unspecified) | |||||
| Nodal segment | Unspecified HgCl2 + Tween-20 for 5 min → thorough rinse with sterile water | MS + 1.0 mg/l BA + 0.05 mg/l NAA | 24 ± 2 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 2000–2500 lux | 10 shoots/explant within 4 weeks with maximum elongation | Wasn’t carried out | [9] |
| MS + 0.5 IBA | Rooting with root length 3.5–4 cm after 3 weeks | |||||
| MS + 0.05 mg/l NAA + 0.1 mg/l BA + 0.1 mg/l Kn | Dark green, compact and hard callus | |||||
| Leaf | Unspecified | MS + 1 mg/l 2,4-D + 0.5 mg/l Kn | 27 ± 2 °C temp, unspecified RH, 16 h photoperiod, with unspecified irradiance | 70% callusing from leaf explants after 4 weeks | Wasn’t carried out | [7] |
| Petiole | MS + 1 mg/l 2,4-D + 1 mg/l Kn | 45% callusing from petiole explants after 4 weeks | ||||
| Friable calli | MS (liquid) + 1 mg/l 2,4-D + 0.5 mg/l Kn | Rotary shaker at 120 ± 5 rpm | Cell suspension culture | |||
| Leaf | Few drops Tween 80 for 15–20 min → 0.1% HgCl2 for 3–4 min | MS + 6 mg/l IAA + 6 mg/l Kn | 25 ± 2 °C temp, 55–6-% RH, 16 h photoperiod, with an irradiance of 2000 lux | Induction and proliferation of friable calli | Wasn’t carried out | [44] |
| Nodal segment | MS + 3 mg/l IBA + 3 mg/l BA | Induction and proliferation of friable calli | ||||
| Root | MS + 3 mg/l IBA + 6 mg/l BA | Induction and proliferation of friable calli | ||||
| Friable calli | PGR-free MS (liquid) | Rotary shaker at 120 rpm | Cell suspension culture | |||
| Shoot tip, Nodal segment | 1% Dettol for 10 min → 0.1% HgCl2 1–4 min → thorough rinse with sterile water | MS + 2 mg/l BA + 0.5 mg/l NAA + 0.5 mg/l TDZ | 25 ± 2 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 1000 lux | 100% of explants produced 23.3 shoots/explant in 28 days | 98.2% plantlets were acclimatized in garden soil + sand + vermicompost (1:1:1) in 28 days | [37] |
| MS + 0.1 IBA | 5.8 roots/shoot with 2.5 cm root length in 17 days | |||||
| Petiole | 0.1% HgCl2 for 5 min → 5 interim rinse for 10 min with sterile water | MS + 0.25 mg/l TDZ + 0.25 mg/l NAA | 25 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 35 μmol/m2/s PPFD | 100% callus induction, 90.6% regeneration with 8.10 shoots per callus | 90% were acclimatized in garden soil within 12 weeks | [31] |
| SH + 0.5 mg/l IBA | 75% rooting with 9–10 roots/ shoot | |||||
| Root | 0.1% HgCl2 for 1–1.5 min → 3 rinse with sterile water | MS + 3.5 mg/l NAA + 1.25 mg/l BA | 25 ± 1 °C temp, unspecified RH, 16 h photoperiod, with an irradiance of 3000 lux | Induction and proliferation of friable calli | Wasn’t carried out | [54] |
| Leaf | 1% Bavistine® solution for 10 min → 1% Savlon 10 min → 70% ethanol for 30 sec → 0.1% HgCl2 for 3 min → 5–7 interim rinse with sterile water | MS + 1 ppm 2,4‑D + 1 ppm BA + 1 ppm IAA | 25 ± 1 °C temp, 70% RH, 16 h photoperiod, with an irradiance of 2000–3000 lux | Profuse growth of soft creamy colored calli | Wasn’t carried out | [40] |
| MS (liquid) + KNO3 + NaCl | Rotary shaker at 120 rpm | Cell suspension culture | ||||
| Nodal segment | 70% alcohol for 1 min → 0.1% HgCl2 (w/v) for 5 min → 4–5 interim rinse with sterile water | MS + 10.0 mg/l BA | 25 ± 2 °C temp, unspecified RH, 12 h photoperiod, with unspecified irradiance | 93.33% explants produced 10.6 shoots with 5.2 cm length | 80% plantlets were acclimatized in garden soil and compost (2:1) | [34] |
| MS + 0.05 mg/l IAA + 0.05 mg/l NAA + 1.0 mg/l BA | 100% induction of light green callus in 14 days | |||||
| MS + 10 mg/l BA | Callus mediated shoot regeneration | |||||
| MS + 1 mg/l IBA | Rooting | |||||
| Nodal segment | 3% (v/v) H2O2 for 2 min → 95% (v/v) ethanol for 1 min → 3 interim rinse with sterile water | MS + 1.1 mg/l BA | 25 ± 1 °C temp, a 16 h photoperiod, with an irradiance of 60 μmol/m2/s PPFD | Shoot initiation in 6 days, 7.4 shoots of 7.2 cm length, 2.8 of leaves/ shoot | Initially in soil and sand (1:1; v/ v) for 4 weeks recording a survival rate of 95%. Finally, plantlets were established in sand, soil and farmyard manure (1:1:1; v/v) for another 4 weeks | [10] |
| MS + 1 mg/l IBA + 0.25 mg/l NAA | 94% rooting, 8.4 roots/shoot with 5.6 cm length | |||||
| MS + 1 mg/l 2,4-D | 46% callus induction that subsequently induced 60 roots per callus, devoid of adventitious shoots | |||||
| Root segment | 0.1% HgCl2 (w/v) for 5 min → 3 interim rinse with sterile water | MS + 1 mg/l 2,4-D + 4 mg/l BA | 25 ± 2 °C temp, 60% RH, unspecified photoperiod, 8 days on rotary shaker (120 rpm) | Cell culture, maximum cell biomass (47.43 g/flask) was achieved | Wasn’t carried out | [39] |
2,4-D 2,4-dichlorophenoxyacetic acid; AC activated charcoal; B5 B5 medium, or Gamborg’s medium [52]; BA N6-benzyladenine; CM coconut milk; CW coconut water; GA3 gibberellin A3; IAA indole-3-acetic acid; IBA indole-3-butyric acid; Kn kinetin or 6-furfurylaminopurine; MS Murashige and Skoog medium [48]; NAA α-napthalene acetic acid; PGR plant growth regulator; Pic piclorum; SH Schenk and Hildebrandt [55]; TDZ thidiazuron.