Figures (5)  Tables (4)

    • Figure 1. 

      (a) Ten-year-old Ilex decidua mother plant, and (b) the explants from epicormic shoots at the lower portion of trunks.

    • Figure 2. 

      The disinfected explants were inoculated on the initiation medium WPM supplemented with (a) 0.5 mg/L BA; (b) explants generated axillary buds three weeks after inoculation; (c) the buds developed into lateral shoots 1−2 cm long in the next two weeks for subculture.

    • Figure 3. 

      Effects of cytokinin type and concentration on the in vitro shoot proliferation in Ilex decidua. (a) Control, no cytokinin; (b) 0.1 mg/L BA; (c) 0.5 mg/L BA; (d) 1.0 mg/L BA; (e) 0.01 mg/L CPPU; (f) 0.1 mg/L CPPU; (g) 0.5 mg/L CPPU; (h) 0.1 mg/L 2-iP; (i) 0.5 mg/L 2-iP; (j) 0.1 mg/L ZT; (k) 0.5 mg/L ZT. (Scale bars = 1 cm, initial subcultured shoots length is about 1 cm).

    • Figure 4. 

      Effects of different basal media with 0.2 mg/L BA and 0.02 mg/L CPPU on in vitro shoot proliferation of Ilex decidua. (a) WPM medium; (b) DKW medium; (c) LP medium; (d) adventitious shoots proliferated from individual explant on LP medium. (Scale bars = 1 cm, initial subcultured shoots length is about 1 cm).

    • Figure 5. 

      Rooting and acclimatizing of Ilex decidua. (a) In vitro rooting on 1/2 LP medium with 0.5 mg/L IBA; (b) ex vitro rooting of micro-cuttings 2 weeks after quick dip in 1 g/L IBA; (c) rooted plantlets transplanted on 10 cm pots with the substrate of 1/2 peat and 1/2 perlite; (d) the plantlets grew to about 20 cm in height two months after transplanting in a greenhouse.

    • Cytokinin Concentration (mg/L) Multiplication rate Shoot length (mm)
      Control 0 1.5 ± 0.13e 5.1 ± 0.5c
      BA 0.1 2.2 ± 0.29de 12 ± 1.5ab
      0.5 6.6 ± 0.55b 10 ± 1.2b
      1.0 9.1 ± 0.81ab 4.2 ± 0.3c
      CPPU 0.01 3.2 ± 0.37cd 16 ± 1.4a
      0.1 9.7 ± 0.84ab 14 ± 1.1a
      0.5 11.6 ± 1.33a 3.2 ± 0.2d
      2-iP 0.1 3.5 ± 0.44cd 4.1 ± 0.5c
      0.5 6.7 ± 0.57b 3.1 ± 0.4d
      ZT 0.1 3.7 ± 0.44c 5.2 ± 0.6c
      0.5 5.6 ± 0.67bc 3.3 ± 0.5d
      Means in each column followed with different superscript letters are significantly different (p ≤ 0.05) according to Duncan's multiple range test.

      Table 1. 

      Effects of cytokinin type (BA, 6-benzylaminopurine; CPPU, N-(2-chloro-4-pyridyl)-N-phenylurea; 2-iP, 6-(γ,γ-Dimethylallylamino)purine; ZT, zeatin) and concentration on the in vitro adventitious shoot multiplication and shoot length of Ilex decidua.

    • BA (mg/L) CPPU (mg/L) Multiplication rate Shoot length (mm)
      0.1 0.01 2.8 ± 0.3d 11 ± 1.5b
      0.2 0.01 6.1 ± 0.4b 13 ± 0.9b
      0.5 0.01 7.6 ± 0.9ab 14 ± 1.6b
      0.1 0.02 4.2 ± 0.5c 21 ± 1.8a
      0.2 0.02 4.7 ± 0.3bc 19 ± 2.2a
      0.5 0.02 8.2 ± 0.7a 9.1 ± 0.7bc
      0.1 0.05 4.3 ± 0.4c 11 ± 1.3b
      0.2 0.05 6.5 ± 0.7b 7.2 ± 0.8c
      0.5 0.05 9.2 ± 0.8a 5.1 ± 0.6d
      Means in each column followed with different superscript letters are significantly different (p ≤ 0.05) according to Duncan's multiple range test.

      Table 2. 

      In vitro multiplication of Ilex decidua adventitious shoots on WPM media with different combinations of BA (6-benzylaminopurine) and CPPU (N-(2-chloro-4-pyridyl)-N-phenylurea).

    • Medium Multiplication rate Average shoot length (mm) Growth observation
      WPM 4.7 ± 0.3a 21 ± 2.5a Small leaves, upward curved, some lower leaves yellow
      MS 1.7 ± 0.2b 18 ± 2.2a Leaves large and thin
      DKW 2.2 ± 0.2b 20 ± 1.6a Leaves large and thin
      LP 5.5 ± 0.4a 22 ± 1.8a Leaves mediate, healthy
      Means in each column followed with different superscript letters are significantly different (p ≤ 0.05) according to Duncan's multiple range test.

      Table 3. 

      Effect of basal medium on in vitro shoot proliferation of Ilex decidua.

    • In vitro rooting Ex vitro rooting
      Auxins (mg/L) Rooted cuttings (%) Number of roots Auxins (g/L, quick dip) Rooted cuttings (%) Number of roots
      0 15 ± 0.8b 4.2 ± 0.3b 0 12 ± 1.1c 3.1 ± 0.2c
      0.5 IBA 100a 9.3 ± 0.5a 0.5 IBA 85 ± 0.6a 7.5 ± 0.6ab
      1.0 IBA 100a 11.2 ± 0.9a 1.0 IBA 100a 9.2 ± 1.2a
      1.5 IBA 100a 12.5 ± 1.4a 1.5 IBA 96 ± 1.1a 8.8 ± 0.8a
      0.5 NAA 90 ± 7.4a 9.6 ± 0.7a 0.5 NAA 77 ± 0.9b 6.6 ± 0.5b
      1.0 NAA 96 ± 8.6a 10.7 ± 0.6a 1.0 NAA 91 ± 0.7a 8.4 ± 0.9a
      1.5 NAA 100a 11.1 ± 1.5a 1.5 NAA 90 ± 0.7a 7.3 ± 0.5ab
      Means in each column followed with different superscript letters are significantly different (p ≤ 0.05) according to Duncan's multiple range test.

      Table 4. 

      Rooting percentage and root number of micropropagated shoots by in vitro and ex vitro rooting with different concentrations of IBA (indole-3-butyric acid) and NAA (α-naphthalene acetic acid) in Ilex decidua.