(Phalaenopsis Silky Moon)

2549

TISSUE CULTURE AND IN VITRO FLOWERING INDUCTION OF PHALAENOPSIS

HYBRID (PHALAENOPSIS SILKY MOON)

By
Tuangporn Rojanawong

A Thesis Submitted in Partial Fulfillment of the Requirements for the Degree

MASTER OF SCIENCE
Department of Biology
Graduate School
SILPAKORN UNIVERSITY
2006


(Phalaenopsis Silky
Moon)

...........................................................
( . )

.............................. ............

1. .
2. .

....................................................
( )
............/......................../..............
....................................................
( . )
............/......................../..............
....................................................
( . )
............/......................../..............

....................................................
(. )
............/......................../..............

46303207 :
: //
:
(Phalaenopsis Silky Moon). :
.. .. . 82 .

(Phalaenopsis Silky Moon) 3 98.65

Hyponex peptone 2 100
1 4
26.23
Calcium hypochlorite [Ca(OCl)2] 7 30

Hyponex peptone 2 6- Benzylaminopurine (BA) 22.2
150
3
protocorm-like bodies (PLBs)
Murashige and Skoog (MS) BA 22.2
PLBs 12.4 3 Hyponex
Saturnfert peptone 2 30
0.5 PLBs 7.9
2
Vacin and Went (VW) 30
40 VW VW 4.5
BA 66.6 30-43
40 0.5-1

2549
................................................
1. ..................................... 2. .....................................

46303207 : MAJOR : BIOLOGY

KEY WORD : PHALAENOPSIS/PROPAGATION/TISSUE CULTURE
TUANGPORN ROJANAWONG : TISSUE CULTURE AND IN VITRO FLOWERING
INDUCTION OF PHALAENOPSIS HYBRID (PHALAENOPSIS SILKY MOON). THESIS
ADVISORS : ASST. PROF. CHOCKPISIT THEPSITHAR, Ph. D. AND ASSOC. PROF. AREE
THONGPUKDEE, Ph. D. 82 pp.
A study on the optimum capsule-age and the best media for seed germination of
Phalaenopsis hybrid (Phalaenopsis Silky Moon) was determined. Seeds from a 3-month-old
capsule provided the highest percentage of germination at 98.65% on liquid modified
Hyponex medium supplemented with 2 g/l peptone, 100 g/l potato juice and 1 g/l activated
charcoal. However, seeds from a 4-month-old capsule showed the highest number of plantlet
regeneration on the same medium. On the basis of flower-stalk bud culture, surfacesterilization with 7% Calcium hypochlorite [Ca(OCl)2] for 30 min. was considered to be the
best pretreatment for decontamination. The solid modified Hyponex medium supplemented
with 2 g/l peptone, 22.2 M BA and 150 ml/l coconut water was shown to be the most suitable
culture medium for flower-stalk bud development. The growth patterns of buds could be
classified into three modes; remaining dormant, growing vegetatively and growing
reproductively depending on the bud position on the stalk. For protocorm-like bodies (PLBs)
induction from leaf segments, a half-strength of Murashige and Skoog (MS) medium
supplemented with 22.2 M 6- Benzylaminopurine (BA), 150 ml/l coconut water and 10 mg/l
adenine sulfate was the best medium provided the highest number of PLBs with 12.4 PLBs
per explant within 3 months. Subsequently the transferred PLBs gave rise to the best plantlet
regeneration at 7.9 plantlets after 2 months on modified Hyponex with Saturnfert fertilizer
containing 2 g/l peptone, 30 g/l potato juice and 0.5 g/l activated charcoal. The induction of
flower buds on Vacin and Went (VW) medium supplemented with 30 and 40 g/l sucrose prior
to transferred to solid VW medium and modified VW medium with 4.5 mM total nitrogen and
66.6 M were added provided the highest percentage of flower bud induction at 30-43%. The
first flower-stalk could be observed after 40 days of culturing and with an average length of
spike 0.5-1 cm.

Department of Biology
Graduate School, Silpakorn University
Academic Year 2006
Student's signature................................................
Thesis Advisors' signature 1.......................................... 2..........................................


.
.

.


....................................................................................................................
...............................................................................................................
.....................................................................................................................
............................................................................................................................
.................................................................................................................................
...........................................................................................................................

1 .............................................................................................................................
1. .............................................................
2. ...................................................
3. ........................................................................................
4. ................................................................................
2 ......................................................................................
1. ................................................................................
2. (Phalaenopsis) ...................................................
2.1 .......................................................................................
2.2 .........................................................................
3. .......................................................................................
4. .....................................................................................
5. .............................................................................
6. .........................................................
7. .................................................................
8. protocorm-like bodies (PLBs) .........................
9. ............................................................................................
10. ..............................................
3 ...............................................................................
1. ...........................................................................................
2. .....................................................................................................

1
1
2
2
3
4
4
4
4
6
8
9
9
12
13
13
15
17
19
19
21

2.1
.................................................................................
2.2
.....................................................................................
2.3 PLBs
2.4 PLBs
..................................................................................................
2.5
................................................
................................................................................................................
1. ...
2.
..............................................................................................
3. PLBs ............
4. PLBs .............
5.
................................................................
..................................................................................................
1.
................................................................................
2. ..........
3. PLBs .......
4. PLBs
5.
......................................................................
........................................................................................................
1.
.............................................................................................................

21
23
25
27
28
30
30
37
44
48
51
57
57
58
59
60
61
63
63


2.
........................................................................................................
3. PLBs ......
4. PLBs .......
5.
......................................................................
..............................................................................................................................
............................................................................................
.............................................................................................................................
..................................................................................................................................
....................................

..............................................................................................................
..............................................................................................................................

63
64
64
64
65
65
66
71
72
77
81

1
2
3
4

5
6
7

8
9

10

11

12

Phalaenopsis Silky Moon.............. 22

Phalaenopsis Silky Moon........................................................................ 22

Phalaenopsis Silky Moon........................................................................ 23

BA NAA
MS 30
Phalaenopsis Silky Moon......................................................................... 25
PLBs Phalaenopsis Silky Moon
.......................................................................... 27
Phalaenopsis Silky Moon .... 28

Phalaenopsis Silky Moon
90 ....................................................................................................... 34
1-5
Phalaenopsis Silky Moon 5 .................................... 38

Phalaenopsis Silky Moon

60 ......................................................................... 41
BA NAA MS
Phalaenopsis Silky Moon
90 ................................................................. 45
PLBs
Phalaenopsis Silky Moon 120
60 ....................................................................................... 49
Phalaenopsis Silky Moon 6 (
2 VW
20, 30, 40 50 ) ...................... 52


13

Phalaenopsis Silky Moon

120 .....................................................................................................

55

1
2
3
4
5
6
7
8
9
10
11

12

13
14
15

(Phalaenopsis) ............................ 5
(Phalaenopsis Silky Moon) .......... 7

(Phalaenopsis Silky Moon) ..................................................................... 8

(flower bud initiation)
(bud development)................................................................................... 16
Phalaenopsis Silky Moon ......................................... 21
Phalaenopsis Silky Moon....................................... 24
Phalaenopsis Silky Moon
PLBs .............................................................. 26
Phalaenopsis Silky Moon
2 ................................................................................ 29
Phalaenopsis Silky Moon
2, 3 4 ........................................... 31
Phalaenopsis Silky Moon
6 ....................................................................................... 32
Phalaenopsis Silky Moon
3 6
90 ..................... 35
Phalaenopsis Silky Moon
4 6
90 .................... 36

Phalaenopsis Silky Moon ......................................................... 39

60 ....................................................................................................... 42
1-5 ................................. 43


16
17

18
19

20

Phalaenopsis Silky Moon..................... 46

Phalaenopsis Silky Moon
0.11
90 MS
BA (0-111.0 ) NAA 0-10.74
) 14 ................................. 47
PLBs
Phalaenopsis Silky Moon 120 ..................................... 50
Phalaenopsis Silky Moon 6
2 VW
4 ............ 53
Phalaenopsis Silky Moon 6
VW BA 66.6
40 ................................................................................ 56

BA
Ca(OCl)2
GA
g/l
MS
mg/l
mM
NAA
PLBs
PVP
TDZ
tTCLs
VW
M

=
=
=
=
=
=
=
=
=
=
=
=
=
=
=

6- Benzylaminopurine
Calcium hypochlorite
Gibberellin
Gram/litre
Murashige and Skoog
Milligram/litre
Millimolar
- Naphthalene acetic acid
protocorm-like bodies
Polyvinylpyrrolidone
Thidiazoron
transverse thin-section culture
Vacin and Went
Micromolar

1.

2549
36.8 1.7
( 2549)
(Genus) (Phalaenopsis) 50
(Species)



75
(Griesbach 2002)
Phalaenopsis Silky Moon

labellum 80

2
(seed propagation) (vegetative propagation)

( 2521)

2
(tissue culture)
( 2546)

Phalaenopsis Silky Moon

2.
1.
2.

3. protocorm like bodies (PLBs)

PLBs
4.

3.
1. (2-4 ) 6
2. 3

3. 30 PLBs
3 PLBs
4.
Vacin and Went, 1949 (VW)

3
4.
1.
Phalaenopsis Silky Moon
2.
3. PLBs

4.
VW

(monocotyledonous plant)
(Family Orchidaceae)
(perennial herbs)
600
(genus) 19,000


( 2516; 2549)
1.




(staminal column) 6 (sepal) 3
(petal) 3

( 2516)
2. (Phalaenopsis)
2.1
(Phalaenopsis) Orchidaceae

5
4
(Phalaenopsis cornucervi (Breda) Bl. & Rchb.f.) ( 1A) (Phalaenopsis lowii Rchb.
f.) ( 1B) (Phalaenopsis parishii Rchb. f.) ( 1C) (Phalaenopsis
decumbens Holtt.) ( 1D)

1 (Phalaenopsis)
(A) (Phalaenopsis cornucervi (Breda) Bl. & Rchb.f.)
(B) (Phalaenopsis lowii Rchb. f.)
(C) (Phalaenopsis parishii Rchb. f.)
(D) (Phalaenopsis decumbens Holtt.)
: ,
( : 2549), 402-405.
: . [Online], Accessed 27 September
1999. Available from http://www.panmai.com/Orchid/Phal/phal.html

6
2.2

(monopodial)







( 2, 3) 6
2 (sepal) 3 (petals) 3 1
3 (mid lobe) 2 (side
lobes)
(column)
(pollinia) 2




(embryo)

1
2-5 ( 2521)



(epiphytic)
( 2546)

7
Phalaenopsis Silky Moon
Phalaenopsis Musashino Phalaenopsis Paper Moon
8-10
80

( 2, 3)

2 (Phalaenopsis Silky Moon)

3 (Phalaenopsis Silky Moon)

LP = lateral petal, DS = dorsal sepal, LS = lateral sepal, ML = mid lobe, SL = side lobe
C = column, AC = anther cap, P = pollinia
3.



( 2516)
3.1
2
3.1.1 (vegetative propagation)

3.1.2 (seed propagation)

4.



( 2546)




5.

Knudson 1921-1922

VW (Knudson 1922; 2536)

10
5.1
5.1.1 (inorganic component) 2
(Arditti and Ernst 1993)
5.1.1.1 (Macroelements)
(Ca) (Mg) (N)
(P) (K) (S)
KNO3, KH2PO4, K2HPO4 KCl
KNO3, NH4NO3, Ca(NO3)2, (NH4)2SO4
5.1.1.2 (Microelements)
(Mn) MnCl2 Mn(NO3)2 MnSO4
(Cu) CuSO4 (Zn) ZnCl2 ZnSO4H2O (B)
H3BO3 (Cl) (Mo) Na2MoO4
MoO3 (S)

EDTA (Ethylene diamine tetraacetic acid) NaFeEDTA
FeSO47H2O (Al) (Co)
(I) (Ni) ( 2546)
5.1.2 (organic component)
5.1.2.1

(Pierik 1987)
(sucrose) (disaccharide)

( 2549; 2536)
5.1.2.2 (amino acid)
(metabolism)

adenine sulphate
(Razdan 2003)
(glycine) Murashige and

11
Skoog 1962 (MS) (Arditti and
Ernst 1993)
5.1.2.3 (vitamins)
(
2549) MS B3 (nicotinic acid),
B6 (pyredoxine) B1 (thiamine) (Arditti and Ernst 1993)
5.1.2.4 (natural complexes)
protein hydrolysates
( casein hydrolysate, peptone tryptone)
( 2540)

(polyamine) biosynthetic enzyme

(nucleic acid) (mitosis)


(Arditti and Ernst 1993)
5.1.2.5 (activated charcoal)

(phenoic componds)
pH ( 2544; Arditti
and Ernst 1993)
5.1.3 (plant growth regulator)

( 2537)


PLBs

12
6.
.. 1899 Bernard
(mycorrhiza) (symbiosis)

Bernard
(Arditti 1967) .. 1922 Lewis Knudson

Knudson B 4 Ca(NO3)2,
K2HPO4, MgSO47H2O, (NH4)2SO4 1 Fe(PO4)3
(asymbiotic method)
Knudson

(2540) Dendrobium Acriopsis

Dendrobium Merritt Island, Dendrobium
Montakan Dendrobium unicum 2
Schenk and Hildebrandt
150 20 CU-1
KNO3 1.25 150
40 Acriopsis indica
CU-1
Alam (2002) Dendrobium transparens
4 Hyponex, MS, OKF1 Knudson C (Knudson, 1964)
MS 78
50 Hyponex 73
55 OKF1
58 59

13
7.

Tanaka Sakanishi (1978b)

6- Benzylaminopurine (BA)
()
20 25
28
BA
BA 2.5
Tanaka (1988)
(8, 16 24 )
0.25 Wm-2 79
Polyvinylpyrrolidone (PVP)
Chen (2003) VW
Gamborg, 1968 (B5) BA 5.0
MS B5, BA 5.0 Naphthalene acetic acid (NAA) 0.5

Hyponex 100 2

8. protocorm-like bodies (PLBs)

rhizoid protocorm
protocorm protocorm-like bodies
(PLBs) (Pierik 1987)

14
(2541) PLBs Phalaenopsis (Phalaenopsis Abendrot
Kelvin) (Kyoto)
NAA 1 adenine 10 BA 10
20 PLBs
Mizono Monma, 1993 MS 15
10 PLBs PLBs
WS 50 50
2 6 PLBs
(2546) Phalaenopsis violacea Witte
PLBs 90
PLBs 46 56 VW Hyponex BA
NAA 5 PLBs
90 PLBs 25.57
VW BA 5 10 PLBs
Hyponex 3
PLBs
Wang (1989) 2
(Kyoto) MS BA 3
MS BA 5
NAA 1 PLBs 17 PLBs
MS BA 1
Myint (2001)
PLBs PLBs VW BA 10
NAA 10 PLBs
VW BA 1.0 NAA 0.1 (agar) 0.8
95 New
Dogashima (ND) BA 1.0 NAA 1.0
30 20 PLBs
PLBs

15
Park (2002) PLBs Doritaenopsis
transverse thin-section culture (tTCLs)
1 MS
adenine sulfate 10 20 (Gelrite) 2.3
3 Thidiazoron (TDZ) (4.522.5 ) BA (2.2-22.2 ) Zeatin (2.2-22.2 )
PLBs 72.3
TDZ 9.0 PLBs 3
9.

(Vegetative Growth)
(Reproductive Growth)
(Ripeness to Flower)



(Growth Regulators)
3
1. (flower bud induction)

(floral primordia) (leaf primordia)
2. (evocation)

RNA

3. (floral development)

16
Phalaenopsis
(monopodial) (bud primordial)

(Rotor 1959) ( 4)

4 (flower bud initiation) (bud

development) A. ; B C.
(floral bud primordia); D, E F.
Am = apical meristem, Br = bract. Fl = floral bud primordia, Se = sepal,
Co = column, La = labellum
: , Rotor, G.B., The Photoperiodic and Temperature
Responses of Orchids. In The orchids : a scientific survey, 400. Edited by Carl L.Withner. New
York: The Ronald press company, 1959.

17

Ichihashi (2003)

140 ppm. 280 ppm.
Kataoka (2004)

(CO2)
(4.3 mol m-2 ) CO2 (1,000-2,000
) 20 2-3

10
10.


(Begonia) (Ringe and Nitsch 1968) (Dendrocalamus hamiltonii Munro.) (Chambers
at al. 1991)
3-13


Duan Yazawa (1995a)
Phalaenopsis Pink Leopard Petra
Hyponex BA VW
BA
BA

18
VW 4.5
BA 5
Duan Yazawa (1995b)
x Doriella Tiny (Doritis pulcherrima x Kingiella philippinensis)
Phalaenopsis Hyponex
BA 5 100
2 PLBs 3-4 3-4
VW BA 5
2 Hyponex BA
peptone
BA
Doriella tiny
(2540)
Dendrobium, Cattleya, Oncidium Ascocenda
Dendrobium Dendrobium bigibbum Dendrobium Pinky Sem
Sabin VW BA 5
40 30
Kostenyuk (1999)
Cymbidium
Cymbidium niveo-marginatum Mak BA 5 10
MS
50 40
TDZ 2.5 75

19

3

1.
1.1
Phalaenopsis Silky Moon
1.2
1.2.1
- MS (Murashige and Skoog, 1962)
- VW (Vacin and Went, 1949)
- Hyponex (N:P:K = 6.5-6-19; Hyponex Co., Marysville, Ohio, USA)
- Saturnfert (N:P:K = 20-20-20; , ,
)
- Phytagel (Sigma, P 8169)
- (activated charcoal)
-
1.2.2
- 6- Benzylaminopurine (BA)
- - Naphthalene acetic acid (NAA)
1.3
- Calcium hypochlorite [Ca(OCl)2]
- Clorox (5.25% w/w Sodium hypochlorite)
- Antibiotic: Streptomycin, Kanker-X, Benlate
- Tween 20

20
1.4
1.4.1
-
- -
-
-
-
-
-
-
-
-
-
1.4.2
- : ,
- : , (scalpel), (forcep)
-
- (larmina flow)

21
2.
2.1
2.1.1

( 5)

5 Phalaenopsis Silky Moon

2.1.2

Phalaenopsis Silky Moon 2, 3 4

10
5 5 70
5 Clorox 30 10
1 3 5
95
1
6 ( 1)

22
1 Phalaenopsis Silky Moon

1 MS
2 Modified.MS
(MMS)
3 VW
4 Modified.VW
(MVW)
5 H
6 Modified.H
(MH)

MS + sucrose (30 g/L) + charcoal (1 g/L)

MS + peptone (2 g/L) + potato juice (100 g/L) + sucrose (30 g/L) +
charcoal (1 g/L)
VW + sucrose (20 g/L) + charcoal (1 g/L)
VW + peptone (2 g/L) + potato juice (100 g/L) + sucrose (20 g/L) +
charcoal (1 g/L)
Hyponex (3.5 g/L) + sucrose (20 g/L) + charcoal (1 g/L)
Hyponex (3.5 g/L) + peptone(2 g/L) + potato juice(100 g/L) +
sucrose(20 g/L) + charcoal (1 g/L)

phytagel 2.5
10 1 pH 5.4 (
1 N KOH 1N HCl) 121
15 30-35
-2 -1
mol.m .s 12
3
6 ( 2)
2 Phalaenopsis Silky Moon

1
2
3
4
5
6

1
1
1

23
Duncans new multiple range test
90
2.2
Phalaenopsis Silky Moon 1-5 ( 6)
1 3
10 5 5
70 5 5
1. Clorox 30 10 Clorox 25
3 3
2. Streptomycin 5 30 Clorox 30
10 3
3. Kanker-X 0.1 25 Benlate 2
10 Clorox 30 10
3
4. Ca(OCl)2 7 30 3

5. 1 30 Ca(OCl)2 7
30 3
3
( 3)
3 Phalaenopsis Silky Moon

1 MS + BA (22.2 M) + coconut water (150 ml/L) + sucrose (30 g/L) + gellite (2 g/L)
2 VW + B5 vitamin (2 ml/L) + BA (22.2 M) + coconut water (150 ml/L) + sucrose
(20 g/L) + gellite (2 g/L)
3 Hyponex (3.5 g/L) + peptone (2 g/L) + BA (22.2 M) + coconut water (150 ml/L) +
sucrose (25 g/L) + gellite (2 g/L)

24
pH 5.4 ( 1 N KOH 1N HCl)
121 15
10 1
30-35 mol.m-2.s-1 12
252
60

Duncans new multiple range test 90

5
4

3
2
1
A

6 Phalaenopsis Silky Moon

(A)
(B) 1-5 ()

25
2.3 PLBs
Phalaenopsis Silky Moon
3 1 ( 7A)
transverse thin-section culture
1-2 ( 7B) MS
2 BA NAA 30 (
4) 30 150 adenine
sulfate 10 phytagel 2.5
4 BA NAA MS
30 Phalaenopsis Silky Moon
NAA (M)
BA (M)
0.0
22.2
44.4
66.6
88.8
111.0

0.54

2.69

5.37

10.74

1
6
11
16
21
26

2
7
12
17
22
27

3
8
13
18
23
28

4
9
14
19
24
29

5
10
15
20
25
30

pH 5.4 ( 1 N KOH 1N HCl)

121 C 15
2 10 2
30-35 mol.m-2.s-1
12 252
90
PLBs PLBs
Duncans new multiple range test 95

26
1.0 cm

7 Phalaenopsis Silky Moon

PLBs
(A) 1
(B) transverse thin-section culture
1-2 MS

27
2.4 PLBs
PLBs Phalaenopsis Silky Moon
BA 22.2 ( 6 3)
120 3 ( 5)
5 PLBs Phalaenopsis Silky Moon

1 VW + peptone (2 g/L) + potato juice (30 g/L) + sucrose (30 g/L) + charcoal (0.5 g/L) +
phytagel (2.5 g/L)
2 MS+ peptone (2 g/L) + potato juice (30 g/L) + sucrose (30 g/L) + charcoal (0.5 g/L) +
phytagel (2.5 g/L)
3 Hyponex 6.5-6-19 (1g/L) + Saturn fert 20-20-20 (1g/L) + peptone (2 g/L)
potato juice (30 g/L) + sucrose (30 g/L) + charcoal (0.5 g/L) + phytagel (2.5 g/L)
pH 5.6 ( 1 N KOH 1N HCl)
121 15
10 1
30-35 mol.m-2.s-1 12
252
60
PLBs PLBs PLBs
Duncans new multiple range test
95

28
2.5

Phalaenopsis Silky Moon

2 ( 8) VW
peptone 2 100 1
20, 30, 40 50 4
2
VW VW BA 22.2, 44.4
66.6 6 ( 6)
6 Phalaenopsis Silky Moon

1
2
3
4
5
6

VW (N = 9 mM) + BA (22.2 M) + sucrose (20 g/L) + phytagel (2.5 g/L)

VW (N = 9 mM) + BA (44.4 M) + sucrose (20 g/L) + phytagel (2.5 g/L)
VW (N = 9 mM) + BA (66.6 M) + sucrose (20 g/L) + phytagel (2.5 g/L)
VW (N = 4.5 mM) + BA (22.2 M) + sucrose (20 g/L) + phytagel (2.5 g/L)
VW (N = 4.5 mM) + BA (44.4 M) + sucrose (20 g/L) + phytagel (2.5 g/L)
VW (N = 4.5 mM) + BA (6.66 M) + sucrose (20 g/L) + phytagel (2.5 g/L)

pH 5.6 ( 1 N KOH 1N HCl)

121 15
10 1 (1 /)
30-35 mol.m-2.s-1 12
252
Hyponex
120

Duncans new
multiple range test 90

29

8 Phalaenopsis Silky Moon

2

30

1.
2, 3 4
6 1

1.1
Phalaenopsis Silky Moon

2, 3 4 8,
8.5 9 2
3 4

(
9)





6 ( 10)
1 =
2 =
3 =
4 = 1
5 = 1
6 = 1

31
1.0 cm

A
1.0 mm.

B
1.0 mm

9 Phalaenopsis Silky Moon 2, 3

4
(A) 2
(B) 3
(C) 4

32

10 Phalaenopsis Silky Moon 6

1 =
2 =
3 =
4 = 1
5 = 1
6 = 1

33
1.2
Phalaenopsis Silky Moon 2, 3 4
6 MS, MMS, VW, MVW, H MH ( 1)
90 2
3
( 1-3)
MH ( 11F) MVW ( 11E) 98.65
98.15 3
( 1-6) MH (
11L) MVW ( 11K) 88.50 86.92 ( 7)
4 (1 -6)
MH ( 12L)
MVW ( 12K) 81.62 76.96
( 5-6)
4 MH ( 12L) MVW ( 12K)
26.23 23.15 ( 7)

34
7 Phalaenopsis Silky Moon 90

1/

1/

MS
MMS
VW
MVW
H
MH
MS
MMS
VW
MVW
H
MH
MS
MMS
VW
MVW
H
MH
MS
MMS
VW
MVW
H
MH

2/ 3/ (%)
1
2
3
4
55.04 4.80 cdef 2.81 0.63 ghi 0.00 0.00 h 0.00 0.00 f
51.38 10.16 efg
5.65 1.48 egf 12.88 2.20 e
0.00 0.00 f
58.04 7.33 cde
1.35 1.00 hi
0.00 0.00 h 0.00 0.00 f
15.12 7.29 klm 16.81 12.13 bcd 65.42 15.24 a
0.54 0.88 f
42.42 2.98 h
5.23 1.21 fg
0.35 0.28 h 0.00 0.00 f
20.92 1.11 ijk 12.42 2.50 b
65.31 1.76 a
0.00 0.00 f
46.62 9.62 fgh
0.35 0.34 i
0.00 0.00 h 0.00 0.00 f
22.73 2.33 ij
3.46 1.61 ghi 19.50 6.33 h 2.54 1.55 e
47.77 2.25 fgh
0.23 0.27 i
0.00 0.00 h 0.00 0.00 f
19.08 11.37 ijkl
0.85 0.79 hi 26.37 6.09 d 29.35 4.98 a
44.38 6.07 gh
1.19 0.48 hi
0.38 0.26 h 0.00 0.00 f
11.35 5.30 lm
2.69 1.57 ghi 39.27 8.20 c 24.92 6.66 b
53.88 8.18 cdef 0.12 0.21 i
0.00 0.00 h 0.00 0.00 f
47.65 26.46 defg 5.65 3.94 efg 8.04 4.80 g 0.00 0.00 f
67.65 4.62 ab
4.27 1.22 fgh 0.00 0.00 h 0.00 0.00 f
14.12 4.76 klm 10.23 4.85 bcd 38.65 4.82 c 10.85 1.41 d
62.00 8.85 bc
5.12 1.48 fg
0.46 0.49 h 0.00 0.00 f
6.31 4.44 ijk
8.85 4.63 cde 44.08 9.72 b 12.73 4.07 d
53.35 4.91 cdef 0.00 0.00 i
0.00 0.00 h 0.00 0.00 f
52.73 5.75 defg 1.04 0.40 hi
0.54 0.34 h 0.19 1.67 f
73.85 8.77 a
1.38 0.96 hi
0.00 0.00 h 0.04 0.10 f
24.62 1.95 i
7.50 1.10 def 10.31 3.10 fg 11.38 0.87 d
61.27 10.70 bcd
1.38 0.54 hi
0.92 0.88 h 0.00 0.00 f
12.58 2.67 klm 11.69 4.70 bc 11.88 1.96 fg 19.23 3.04 c

5
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.46 0.44 e
0.00 0.00 e
11.35 2.00 c
0.00 0.00 e
10.00 2.47 d
0.00 0.00 e
0.00 0.00 e
0.00 0.00 e
0.19 0.17 e
0.00 0.00 e
0.92 0.48 e
0.00 0.00 e
0.04 0.10 e
0.00 0.00 e
15.92 2.67 b
0.00 0.00 e
19.31 2.01 a

6
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
0.12 0.21 c
0.00 0.00 c
0.27 0.42 c
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
1.81 1.01 b
0.00 0.00 c
1.73 1.05 b
0.00 0.00 c
0.00 0.00 c
0.00 0.00 c
7.23 1.18 a
0.00 0.00 c
6.92 1.76 a

3/
(%)
57.85 1.79 fghi
69.92 4.22 cdef
59.38 2.85 fghi
98.15 12.69 a
48.00 1.16 hi
98.65 1.40 a
46.96 3.75 hi
48.69 3.80 hi
48.00 0.92 hi
86.92 8.15 ab
45.96 2.40 i
88.50 6.84 ab
54.00 3.10 ghi
61.35 13.09 efgh
71.92 1.95 cdef
75.85 4.20 bcd
67.58 3.84 cedfg
74.62 6.04 bcde
53.35 1.89 ghi
54.54 2.09 ghi
75.27 3.46 bcde
76.96 2.35 bcd
63.58 4.18 defg
81.62 3.49 bc

MS = Murashige and Skoog, MMS = Modified Murashige and Skoog, VW = Vacin and Went, MVW = Modified Vacin and Went, H = Hyponex, MH = Modified Hyponex
1 = , 2 = , 3 = , 4 = 1 , 5 = 1 , 6 = 1
3/
( S.E.) 90 (P = 0.1) DMRT 10
1
2/

35

11 Phalaenopsis Silky Moon

3 6
90
(A) MS (B) VW
(C) H
(D) MMS (E) MVW
(F) MH
(G) MS
(H) VW
(I) H
(J) MMS (K) MVW
(L) MH

36

12 Phalaenopsis Silky Moon

4 6
90
(A) MS (B) VW
(C) H
(D) MMS (E) MVW
(F) MH
(G) MS
(H) VW
(I) H
(J) MMS (K) MVW
(L) MH

37
2.

Phalaenopsis Silky Moon 1-5
1 2-5
( 6) 1 3
5 3 3
2
2.1

( 13) Ca(OCl)2 7
30
1-5 16.7, 30.0, 63.7, 60.0 50.0 Ca(OCl)2
1
2 1-5 10.0, 62.5, 44.4, 50.0 40.0
Streptomycin 5 30
Clorox 30 10
1, 2 4 100 3 5 10.0
30.0 1
( 8)

38
8 1-5 Phalaenopsis
Silky Moon 5

1. 30% Clorox 10 , 25% 1

10
Clorox 30
2
8
3
9
4
12
5
11
2. 5% Streptomycin 30 , 1
8
30% Clorox 10
2
10
3
10
4
10
5
10
3. 0.1% Kanker-X 25 , 1
16
2% Benlate 10 , 30%
2
12
Clorox 10
3
11
4
12
5
12
1
6
4. 7% CaOCl2 30
2
10
3
11
4
10
5
8
5. 1% 30 ,
1
10
7% CaOCl2 30
2
8
3
9
4
8
5
5

(%)
0.0
37.5
11.1
33.3
27.3
0.0
0.0
10.0
0.0
30.0
0.0
0.0
18.2
25.0
16.7
16.7
30.0
63.7
60.0
50.0
10.0
62.5
44.4
50.0
40.0

80.0
20.0
62.5
50.0
37.5
25.0
40.0
33.3
40.0
42.9
93.8
66.7
66.7
44.4
100.0
100.0
71.4
75.0
100.0
100.0
66.7
100.0
80.0
100.0
100.0

(%)

+
10.0 10.0
60.0 20.0
37.5 0.0
50.0 0.0
62.5 0.0
62.5 12.5
40.0 20.0
44.4 22.2
50.0 10.0
14.3 42.9
0.0
6.2
25.0 8.3
33.3 0.0
44.4 11.1
0.0
0.0
0.0
0.0
14.3 14.3
25.0 0.0
0.0
0.0
0.0
0.0
22.2 0.0
0.0
0.0
20.0 0.0
0.0
0.0
0.0
0.0

39
1.0 cm

A
1.0 cm

B
1.0 cm

13
Phalaenopsis Silky Moon
(A)
(B)
(C)

40
2.2
1-5
3
( 14)
3 ( 3) 60
Hyponex ( 15A)
1-5 1.65, 2.35, 3.39, 3.41 3.61
1.4, 1.1, 1.0, 0.8 1.4
VW ( 15B) 1-5 0.74, 1.65,
2.19, 2.34 2.72 0.5, 1.2, 0.2, 0.3 0.2
MS ( 15C)
1-5 0.41, 1.11, 1.21, 1.67 2.47
0, 0, 0, 0.1 0.3 ( 9)

41
9
Phalaenopsis Silky Moon 60
1/

2/
(.)

MS

0.43 0.12 g
1.11 0.15 efg
1.21 0.14 ef
1.67 0.24 de
2.47 0.22 c
0.74 0.18 fg
1.65 0.21 de
2.19 0.12 cd
2.34 0.28 cd
2.72 0.34 bc
1.65 0.46 de
2.35 0.28 cd
3.39 0.41 ab
3.41 0.41 ab
3.61 0.38 a

VW

1/
2/

1
2
3
4
5
1
2
3
4
5
1
2
3
4
5

9
2
0
7
0
6
2
0
0
0
6
1
0
0
0

1
8
10
0
0
4
8
7
0
0
4
9
6
0
0

0
0
0
3
10
0
0
3
10
10
0
0
4
10
10

0.0 0.00 d
0.0 0.00 d
0.0 0.00 d
0.1 0.10 d
0.3 0.15 cd
0.5 0.27 bcd
1.2 0.36 ab
0.2 0.13 d
0.3 0.30 cd
0.2 0.20 d
1.4 0.58 a
1.1 0.23 ab
1.0 0.42 abc
0.8 0.36 abcd
1.4 0.43 a

MS = Murashige and Skoog, VW = Vacin and Went, H = Hyponex,

( S.E.) 90
(P = 0.1) DMRT 10 1

42
1.0 cm

A
1.0 cm

B
1.0 cm

14 60
(A)
(B)
(C)

43

5
1.0 cm

15 1-5
(A) 1-5 Hyponex
(B) 1-5 VW
(C) 1-5 MS

44
3. PLBs
Phalaenopsis Silky Moon
0.11 tTCLs
MS BA 0-111.0 NAA
0-10.74 30 ( 4)
10
( 16A)
14 45 PLBs
( 16C)
90 MS
( 1) 95
BA 22.2 ( 6) 85
BA 66.6, 88.8
NAA 10.74 ( 20, 25) BA
111.0 NAA 2.67 ( 28-30)
( 10)
PLBs MS BA
22.2 ( 6) PLBs 65
PLBs 12.4 BA 44.4
NAA 0.54 PLBs 45
PLBs 5.44 PLBs
BA 66.6 NAA 2.67-10.74
BA 88.8 ( 18-30)
NAA 0.54
BA 22.2 NAA 0.54 ( 2
7) ( 10, 17)

45
10 BA NAA MS
Phalaenopsis Silky Moon
90
1/
PLBs

PLBs (%)
95.0 0.05 a
5.0 0.05 ef
1.00 0.05 c
75.0 0.10 abc 25.0 0.10 cde
1.80 0.20 c
65.0 0.11 bc 10.0 0.07 def
1.50 0.11 c
65.0 0.11 bc 10.0 0.79 def
2.00 0.16 c
55.0 0.11 bc
5.0 0.05 ef
3.00 0.15 c
85.0 0.08 ab 65.0 0.11 a
12.40 1.58 a
75.0 0.10 abc 45.0 0.11 abc
2.22 0.36 c
65.0 0.11 bc 40.0 0.11 bc
2.13 0.26 c
50.0 0.12 cd 25.0 0.10 cde
3.40 0.36 c
65.0 0.11 bc 55.0 0.11 ab
5.36 0.84 b
60.0 0.11 bc 40.0 0.11 bc
2.25 0.30 c
65.0 0.11 bc 45.0 0.11 abc
5.44 0.92 b
65.0 0.11 bc 40.0 0.11 bc
2.00 0.30 c
60.0 0.11 bc 30.0 0.11cd
2.33 0.30 c
45.0 0.11 cde 10.0 0.07 def
1.50 0.11 c
20.0 0.09 ef 10.0 0.07 def
2.00 0.16 c
25.0 0.10 def 15.0 0.08 def
3.00 0.31 c
20.0 0.09 ef
0.0 0.00 f
0.00 0.00 c
20.0 0.09 ef
0.0 0.00 f
0.00 0.00 c
0.0 0.09 f
0.0 0.00 f
0.00 0.00 c
10.0 0.07 f
0.0 0.00 f
0.00 0.00 c
25.0 0.10 def 0.0 0.00 f
0.00 0.00 c
15.0 0.08 f
0.0 0.00 f
0.00 0.00 c
5.0 0.05 f
0.0 0.00 f
0.00 0.00 c
0.0 0.00 f
0.0 0.00 f
0.00 0.00 c
10.0 0.07 f
0.0 0.00 f
0.00 0.00 c
15.0 0.08 f
0.0 0.00 f
0.00 0.00 c
0.0 0.00 f
0.0 0.00 f
0.00 0.00 c
0.0 0.00 f
0.0 0.00 f
0.00 0.00 c
0.0 0.00 f
0.0 0.00 f
0.00 0.00 c

(M)
BA
NAA
(%)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
1/

0.0

22.2

44.4

66.6

88.8

111.0

0.00
0.54
2.69
5.37
10.74
0.00
0.54
2.69
5.37
10.74
0.00
0.54
2.69
5.37
10.74
0.00
0.54
2.69
5.37
10.74
0.00
0.54
2.69
5.37
10.74
0.00
0.54
2.69
5.37
10.74

2/
+++
+
++
+
+
+++
+
++
+
+
+
+
++
+
-

( S.E.) 95
(P = 0.05) DMRT 2 10 20
2/
3
-
+
++
+++

46

1.0 mm

1.0 mm
PLBs

C
1.0 mm

16 Phalaenopsis Silky Moon

(A) 7
(B) 15
(C) 45 C = callus,
PLBs = protocorm-like bodies

47
NAA
BA

0.0 M

0.54 M

2.69 M

10.74 M

5.37 M

0.0 M
1

10

11

12

13

14

15

16

17

18

19

20

22.2 M

44.4 M

111.0 M
17 Phalaenopsis Silky Moon 0.11 90
MS BA (0-111.0 ) NAA (0-10.74 ) 14

48
4. PLBs
PLBs Phalaenopsis Silky
Moon BA 22.2 ( 6 3)
120 ( 18A) 3 VW , MS
Hyponex (6.5-6-19) 1 Saturnfert (20-20-20) 1
peptone 2 100 1
30 phytagel 2.5 PLBs
60 PLBs 100
VW Hyponex
MS 10 ( 11)
PLBs VW ( 18C) PLBs
74 PLBs
PLBs Hyponex (
18D) PLBs 51 PLBs
( 18E) PLBs 7.9
PLBs MS ( 18B)
PLBs 10 PLBs
( 11)

49
11 PLBs Phalaenopsis
Silky Moon 120 60

Modified MS
Modified VW
Modified Hyponex
1/

PLBs1/
(%) PLBs (%)2/ ()
10.0 0.10 b
10.0 1.00 c
0.0 0.00 b
100.0 0.00 a
74.0 3.10 a
0.0 0.00 b
100.0 0.00 a
51.0 5.26 b
7.9 1.35 a

( S.E.) 95
(P = 0.05) DMRT 10 1 PLBs
2/
PLBs PLBs PLBs

50
1 mm.

2 mm.

2 mm.

18 PLBs Phalaenopsis
Silky Moon 120
(A) PLBs MS BA
22.2 ( 6 3)
(B) PLBs MS
(C) PLBs VW
(D) PLBs Hyponex
Saturnfert
(E) 1 PLBs

51
5.

Phalaenopsis Silky Moon

VW 20-50
4
BA
6
( 6)
5.1
VW peptone 2
100 1
4 20, 30, 40 50 40
0.88 3.15
2.27 50
7.35 1.94
( 12, 19)

52
12 Phalaenopsis Silky Moon 6 (
2 VW 20,
30, 40 50 )

(/) (.)
20
0.71 0.04 b
30
0.75 0.05 b
40
0.88 0.04 a
50
0.67 0.03 b

1/

1/
(.) (.)
3.00 0.13 a 2.22 0.11 a
4.35 0.31 b
1.81 0.12 a
3.05 0.14 a 2.00 0.08 a
5.00 0.33 b
1.90 0.13 a
3.15 0.13 a 2.27 0.10 a
7.35 0.43 a
1.94 0.12 a
3.00 0.07 a 1.68 0.10 b
7.70 0.25 a
1.99 0.13 a

( S.E.) 95
(P = 0.05) DMRT 20 1

53

19 Phalaenopsis Silky Moon 6

2 VW
4
(A) VW 20
(B) VW 30
(C) VW 40
(D) VW 50

54
5.2 BA
VW
20-50 4
VW
VW 9 4.5 2 BA
22.2, 44.4 66.6 VW
BA 66.6 40%
VW BA
38 43 VW
30 40 VW
BA 66.6 0.5-1
( 13, 20)

55
13 Phalaenopsis Silky Moon 120

VW +
(/)

1/

20
30
40
50
20
30
40
50
20
30
40
50
20
30
40
50
20
30
40
50
20
30
40
50

BA
1/
(M)

VW
VW
VW
VW
VW
VW
VW
VW
VW
VW
VW
VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW
4.5N-VW

22.2
22.2
22.2
22.2
44.4
44.4
44.4
44.4
66.6
66.6
66.6
66.6
22.2
22.2
22.2
22.2
44.4
44.4
44.4
44.4
66.6
66.6
66.6
66.6

0
0
0
0
0
0
0
0
0
60-120
0
0
120
0
0
0
80
80
100
0
0
60-120
40-120
0

2/

(%)

0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
0.00.00 b
40.00.25 a
0.00.00 b
0.00.00 b
10.00.10 b
0.00.00 b
0.00.00 b
0.00.00 b
14.00.14 b
17.00.17 b
14.00.14 b
0.00.00 b
0.00.00 b
43.00.20 a
38.00.18 a
0.00.00 b

3/

(.)

0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
1.00
0.00
0.00
0.80
0.00
0.00
0.00
1.00
0.50
1.00
0.00
0.00
0.83
1.00
0.00

(.)
(.)

1.340.15
1.360.07
1.080.10
1.010.10
1.030.03
0.860.04
0.850.08
0.910.07
1.300.06
1.260.11
1.040.04
0.960.04
1.130.04
1.410.06
1.280.09
1.110.08
1.360.09
1.450.05
1.360.09
1.320.09
1.340.13
1.200.08
1.310.09
1.150.08

6.30.64
5.10.55
5.10.61
5.40.62
5.10.40
4.70.18
4.40.50
4.40.41
6.00.30
4.40.75
4.60.43
3.40.30
6.40.54
6.30.75
6.00.66
6.60.43
5.90.63
6.70.42
7.10.51
5.60.43
6.50.76
5.30.52
5.00.27
5.00.60

1.550.08
2.390.35
1.950.20
1.950.22
1.650.07
1.790.12
1.910.15
1.720.13
2.040.02
1.810.22
1.700.20
1.650.12
1.730.11
2.290.24
1.900.16
1.570.10
1.650.09
1.510.16
1.410.12
1.350.10
1.540.11
1.720.21
1.630.10
1.570.19

1.00.22
1.30.25
0.30.17
0.50.17
0.40.20
0.70.18
0.50.19
0.40.18
0.00.00
0.00.00
0.00.00
0.00.00
0.40.16
0.50.19
0.80.16
0.90.14
0.40.20
0.50.22
0.40.20
0.60.16
0.00.00
0.00.00
0.00.00
0.00.00

(.)

0.510.11
0.540.29
0.480.26
0.400.15
0.210.10
0.430.13
0.250.09
0.290.12
0.000.00
0.000.00
0.000.00
0.000.00
0.200.08
0.380.16
0.630.18
0.860.14
0.240.12
0.370.18
0.290.15
0.420.13
0.000.00
0.000.00
0.000.00
0.000.00

VW = Vacin and Went, 4.5N-VW = Vacin and Went

( S.E.) 90 (P = 0.1) DMRT
3/

2/

0.000.00
0.000.00
0.000.00
0.000.00
0.140.14
0.000.00
0.130.13
0.000.00
0.000.00
0.000.00
0.000.00
0.000.00
0.200.13
0.130.13
0.250.16
0.290.18
0.290.18
0.000.00
0.290.18
0.200.13
0.500.27
0.000.00
0.000.00
0.000.00

56
1.0 cm.
Fs

A
Br

1.0 mm.

Br

Br
Se

Br

Am

Co La

Fl
Br

B
20 Phalaenopsis Silky Moon 6
VW BA 66.6 40
(A) Phalaenopsis Silky Moon
Fs = floral stalk
(B) (L.S.) A
Am = apical meristem, Br = bract, Co = colum, F = flower, Fl = floral bud
primordial, Se = sepal, La = labellum

57

1.

Phalaenopsis Silky Moon Phalaenopsis Silky Moon

2, 3 4


(carotenoid) ( 2525)
3
2


( 2538) 3


VW (MVW) H (MH)
100 peptone 2 1
peptone Thomale GD
Paphiopedilum (Flame 1978) peptone protein
hydrolysate (amino acids) (Arditti 1977)
arginine

Phalaenopsis

58
(Tsai 1992) (Polyamine)
(nucleic acid)
(mitosis)



(Arditti Ernst 1984)

VW H VW H
MS Arditti
Harison (1977)

2.
Phalaenopsis Silky Moon

Ca(OCl)2 7 30
Tanaka Sakanishi (1978b) Ca(OCl)2 7
10 Phalaenopsis
amabilis 1

Hyponex peptone 2 BA 22.2 150
Phalaenopsis Pink
Leopard Petra (Duan Yazawa 1995a) BA
( 2537)
BA Phalaenopsis
(Tanaka Sakanishi 1978b; Chen 2003; 2521)

59
1-5
3
2 3
4 5 (2521)
flowering hormone

3. PLBs
Phalaenopsis Silky Moon
3
1 0.11
transverse thin-section culture (tTCLs) PLBs
Aranda Deborah (Lakshmanan 1995)
Doritaenopsis (Park 2002)
MS Phalaenopsis 5
MS, VW, KC, LM Hyponex MS
PLBs (Park 2002) 2
Hsu Chen (2003)
PLBs
BA NAA BA

( 2537) NAA
DNA ( 2540)
BA 22.2 adenine sulfate 10
150 PLBs
PLBs (12.4 PLBs ) Duan Yasawa
(1995b) BA 22.2 PLBs
Phalaenopsis (Doriella Tiny) Park (2002) BA 88.8
NAA 5.4 MS
PLBs (12 PLBs ) Phalaenopsis Taisuco

60
Hatarot

4. PLBs
PLBs 3
3 VW, MS Hyponex (6.5-6-19) 1
Saturnfert (20-20-20) 1 peptone 2
30 0.5 PLBs
VW PLBs
VW 20
PLBs (Tanaka Sakanishi 1978a) PLBs

VW (Vacin Went 1949) MnSO4
PLBs PLBs
Hyponex Saturnfert PLBs
PLBs
PLBs 7.9 Park (2002)
Hyponex (6.5-4.5-19 1 + 20-20-20 1 )
peptone 2 3 0.05
30 PLBs Phalaenopsis 4
6 Hyponex
PLBs Phalaenopsis

61
5.

Phalaenopsis Silky Moon

2 Vacin and Went (VW)
100 peptone 2 4
20-50 4
VW VW 9 4.5
BA 3 22.2-66.6
VW
20-50
4 40
50

BA

VW 40 VW BA
66.6 VW
BA
BA
VW 40
(2540)
BA
BA

BA Duan
Yazawa (1995b) x Doriella Tiny (Doritis
pulcherrima x Kingiella philippinensis) VW
BA 5 Duan Yazawa
Phalaenopsis Pink Leopard Petra

62
70 VW
9 4.5 BA 5 Ziv
Naor (2006) 2


Chen
(1994) Phalaenopsis
CO2

63

1.
Phalaenopsis Silky Moon
2, 3 4
6
1.1 Phalaenopsis Silky Moon

3

1.2 Phalaenopsis Silky Moon

1.3 VW
(MVW) H (MH) peptone

2.
Phalaenopsis Silky Moon
Ca(OCl)2 7 30
3 MS, VW Hyponex
Hyponex peptone, BA
3
2 3
4 5

64
3. PLBs
PLBs transverse thin-section culture
(tTCLs) MS BA 0-111.0
NAA 0-10.74 PLBs BA
22.2 adenine sulfate 10 150
PLBs PLBs
12.4 PLBs 3
4. PLBs
PLBs 3
3 VW, MS Hyponex (6.5-6-19) 1
saturn fert (20-20-20) 1 peptone 2
100 1 PLBs
VW PLBs
Hyponex
PLBs 2
5.

Phalaenopsis Silky Moon

VW 30 40
VW VW 9.0 4.5
BA 66.6
30-43
40 0.5-1

65

1.
Hyponex
2.

3. PLBs
PLBs

4. PLBs PLBs
VW PLBs Hyponex

5.
Gibberellin (GA), TDZ

66

. . :
, 2546.
. . 2. :
, 2544.
. .
, 2536.
. 512 521 .
, 2549.
. .
, 2540.
. Phalaenopsis violacea Witte .

, 2546.
. Phalaenopsis Adendrot Kelvin
. (. .) , 2541.
. :
. . 302 (2546): 26-28.
. .
: , 2537.
. . : ,
2516.
. : . :
, 2540.
. [Online] Accessed 27 September 1999. Available from
http://www.panmai.com/Orchid/Phal/phal.shtml

67
. 421553 Plant Morphogenesis.
, 2540.
. : [Online]. Accessed 8 October
2549. Available from http://www.positioning.com/prnews/prnews.aspx?id=50991
. .

, 2538.
. . : , 2525
. . 11. :
, 2549.
. .
. (2521) : 69-80.

Alam, M.K., Rashid, M.H., Hossain, M.S., Salam, M.A. and Rouf, M.A. In vitro seed
propagation of Dendrobium (Dendrobium transparens) orchid as influenced by different
media. Biotechnology. 1(2002): 111-115.
Arditti, J. Factors affecting the germination of orchid seeds. The Botanical Review 3 (1967):
1-20.
Arditti, J., Clements, M.A., Fast, G., Hadley, G., Nishimura, G. and Ernst, R. Orchid seed
germination and seedling culture. Orchid Biology: Reviews and Perspectives, II.
(1982): 266.
Arditti, J. and Harison, C.R. Vitamin requirements and metabolism in orchid. In Orchid
Biology: Reviews and Perspectives I, 159-175. Edited by Joseph Arditti. Ithaca: Cornell
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and Perspectives III, 178-222. Edited by Joseph Arditti. Ithaca: Cornell University Press,
1984.
_______. b Micropropagation of Orchids. New York : John Wiley &Sons, Inc., 1993.

68
Chamber, S. M., Heuch, J.H. R. and Pirrie, A. Micropropagation and in vitro flowering of the
bamboo Dendrocalamus hamiltonii Munro. Plant Cell Tissue and Organ Culture.
27(1991) : 45-48.
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influence carbohydrate content and flowering in Phalaenopsis. Physiologia Plantarum.
90(1994) 391-395.
Chen, Z., Ye, X. and Liang, C. In vitro culture of the inflorescence of Phalaenopsis. Acta
Horticulturae Sinica. 30(2003): 242-244.
Duan, J.X. and Yazawa, S. a Floral induction and development in Phalaenopsis in vitro.
Plant Cell Tissue and Organ Culture. 43(1995): 71-74.
_______. b Induction of precocious flowering and seed from of xDoriella Tiny
(Doritis pulcherrima x Kingiella philippinensis) In vitro and In vivo. Acta Horticulturae
Sinica. 397(1995): 103-110.
Flame, M. Influence of selected media of supplements on the germination and growth of
Paphiopedilum seedling. American Orchid Society Bulletin. 47(1978): 419-423.
Griesbach, R.J. Development of Phalaenopsis Orchids for the Mass-Market. Trend in new
crop and new uses. (2002): 458-465.
Hsu, C. and Chen, F. Plant regeneration from protocorm-like bodies induced in etiolated
leaves of Phalaenopsis Aphrodite Rchb. f.. Journal of the Chinese Society for
Horticultural Science. 49(2003): 335-342.
Ichihashi, S. Effects of nitrogen application on leaf growth, inflorescence development and
flowering in Phalaenopsis. The Bulletin of Aichi University. 52(2003): 35-42.
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Phalaenopsis leaves before floral transition. Scientia Horticulture. 102(2004): 121-132.
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Orchid Society Bulletin. 15(1964): 214-217.

69
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protocorm-like bodies from leaf subepidermal cell of Doritaenopsis hybrid using thinsection culture. Plant Cell Reports. 21(2002): 46-51.
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70
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6(2006): 3-16.

71

72

73
1 Vacin and Went (1949) VW
9 4.5

Ca3PO4
KNO3
KH2PO4
MgSO47H2O
(NH4)2SO4
KCl

FeSO47H2O
MnSO44H2O
Na2-EDTA

Sucrose
phytagel

()
VW
4.5VW
200
525
250
250
500
-

200
262.5
250
250
250
193.7

28.0
7.5
73.0

28.0
7.5
73.0

20,000 (20 g/l)

2,500 (2.5 g/l)

20,000 (20 g/l)

2,500 (2.5 g/l)

74
2 Murashige and Skoog (1962)

MgSO47H2O
CaCl22H2O
KNO3
NH4NO3
KH2PO4

FeSO47H2O
MnSO44H2O
KI
CoCl26H2O
ZnSO47H2O
CuSO45H2O
H3BO3
Na2MoO42H2O

Na2EDTA
Thiamine-HCl
Nicotinic acid
Pyridoxine-HCl
Glycine
Myo-inosital
Sucrose
phytagel

370
440
1,900
1,650
170
27.8
22.3
0.83
0.025
8.6
0.025
6.2
0.25
37.3
0.1
0.5
0.5
2
100
30,000 (30 g/l)
2,500 (2.5 g/l)

75
VW 1
- (NH4)2SO4 132.154 N = 28.02
1,000 (NH4)2SO4 500
N = (50028.02)/132.154
= 106.013
N (NH4)2SO4 = 106.013/28.02 = 3.8
- KNO3 101.11 N = 14.01
1,000 KNO3 525
N = (52514.01)/101.11
= 72.745
N KNO3 = 72.745/14.01 = 5.2
VW = 3.8 + 5.2 = 9

76

1. BA (6- Benzylaminopurine) 1.0 mg/ml

BA 100 0.1% KOH 1-2 ()
1,000
2. NAA ( - Naphthalene acetic acid) 1.0 mg/ml
NAA 100 0.1% KOH 1-2 ()
1,000

77

78
1.

1. (fixative)
Formalin-Aceto-Alcohol (FAA)
50% ( 70%) ethanol
90 .
glacial acetic acid
5 .
formalin
5 .
2. (dehydrating agent)
ethanol n-butanol
3. (embedding media)
(paraplast)

1. 5-8 .

2. FAA 18
3. - 50% ethanol 2-3
- 50% ethanol 2 1-2
- 70% ethanol
4. (dehydration) ethanol n-butanol
(1-5) 50%, 70%, 85%, 95% 100%
1
(1-5):
1.
50 .
95% ethanol
40 .
butanol
10 .
2.
30 .
95% ethanol
50 .
butanol
20 .

79
3.
95% ethanol
butanol
4. 95% ethanol
butanol
5. absolute ethanol
butanol

15 .
50 .
35 .
45 .
55 .
25 .
75 .

5. absolute ethanol 1
6. absolute ethanol eosin light green (

section )
7. ( 58)
1 (P1) 2 (P2, P3 )
1-2 ()
8. (embedding mold)

label
9. section 10-12
M section
10. ( 56-60) 45-60 section

80
2. safranin & fast green
:
1% aqueous safranin
safranin O (C.I. 50240)
1

100 .
0.2% fast green
fast green FCF (C.I. 42053)
0.2
absolute ethanol
100 .
:
1. xylene 2 5-10
2. :
- 95% ethanol 2 3-5
- 70% ethanol 2-3
- 50% ethanol 2-3
-
3. section 1% aqueous safranin O 30-50
4. 2
5. 50%, 70% 95% ethanol 2-3
6. counterstain 0.2% fast green (1-15 )
7. absolute ethanol 2
8. xylene 2 3-5
9. DPX
10. label

81

.. 2542
.. 2545

.. 2546

240 () 101
10240
6

(Phalaenopsis Silky
Moon)

Rojanawong T, Thepsithar C, Thongpukdee A (2006) In vitro flower bud induction of

Phalaenopsis Cygnus Silky Moon from seed-derived plantlets. Proceeding: 32nd
Congress on Science and Technology of Thailand Science and Technology for
Sufficiency Economy to Celebrate the 60th Anniversary of the Majesty the Kings
Accession to the Throne. Sirikit Convention Hall, Bangkok, Thailand, 10-12 October,
F_0008
Rojanawong T, Thepsithar C, Thongpukdee A (2006) In vitro seed culture of Phalaenopsis
hybrids (Phalaenopsis Cygnus Silky Moon). The Sixth National Horticulture Congress,
Lotus Hotel Pang Suan Kaew, Chaingmai, Thailand, 7-10 November, (Abstract), P215, p
329

82
Rojanawong T, Thepsithar C, Thongpukdee A (2006) Micropropagation of Phalaenopsis
Cygnus Silky Moon from leaf segments. Proceeding: 32nd Congress on Science and
Technology of Thailand Science and Technology for Sufficiency Economy to Celebrate
the 60th Anniversary of the Majesty the Kings Accession to the Throne. Sirikit
Convention Hall, Bangkok, Thailand, 10-12 October, F_0006
Rojanawong T, Channuntapipat C, Boonyakitjinda V (2003) Effect of vinegar vapor on
postharvest of papaya. 2nd National Technical Seminar on Postharvest/Post Production
Technology. KhonKhan, Thailand, p 88