EFFECTS OF SEVERAL COMMERCIALLY AVAILABLE SOLIDIFYING ...

Table 3. Continued.

mg B/6 inch

pot/3 trt

Schefflera arboricola

PPM B Plant

in foliage grade2

mg B/4 inch

pot/3 trt

Araucaria heterophylla

PPMB

in foliage

No. necrotic

tips

0

15

30

45

60

35

240

383

397

555

4.3

4.3

3.7

3.3

2.0

0

5

10

15

20

16

107

146

284

564

0.0

0.0

0.0

0.0

10.0

Alpinia zerumbet' Variegata'

mg B/6 inch PPM B No. necrotic

pot/3 trt in foliage leaves

Philodendron scandens oxycardium

mgB/8inch PPMB

pot/4 trt in foliage

Severity of

0

15

30

45

60

37

122

179

385

564

0.0

1.0

0.8

2.5

6.8

0

40

80

120

160

64

175

256

309

487

1.0

1.3

1.8

2.3

4.3

Neoregelia carolinae Tricolor*

mg B/6 inch PPMB

pot/4 trt in foliage

Severity of mg B/8 inch

pot/5 trt

Ficus benjamina

PPMB

in foliage

Severity of

0

20

40

60

80

43

434

645

784

899

1.5

2.5

3.6

3.8

4.5

0

50

100

150

200

67

267

309

410

425

1.0

1.0

1.2

1.4

1.7

zPlants were graded on a scale of 1 = poor quality, unsalable, 3 ySeverity of necrosis was based on a scale of 1 = no necrosis, 3 =

= fair quality, salable and 5 = excellent quality plant material.

■■ some necrosis, but still salable and 5 = severe necrosis, unsalable plant material.

(Poole and Conover, 1982). To avoid B toxicity problems

foliage producers should know the concentration of B in their irrigation water, growing medium, fertilizers and

amendments. If one irreplaceable component of a produc

tion program contains high levels of B, other components

should contain little or no B.

Literature Cited

ben Jaacov, J., R. T. Poole, and C. A. Conover. 1984. Tipburn of C.

comosum 'Vittatum.' HortScience 19(3):445-447. Conover, C. A. and R. T. Poole. 1973. Factors influencing micronutrient

use in tropical foliage production. Univ. Fla. Agr. Res. Center - Apopka

MimeoRpt. RH-73-1.

Kohl, H. C. and J. J. Oertli. 1961. Distribution of boron in leaves. Plant

Physiol. 26(4):420-424.

Marlatt, R. B. 1978. Boron deficiency and toxicity symptoms in Ficus

elastica 'Decora' and Chrysalidocarpus lutescens. HortScience 13(4):442-

443.

Poole, R. T. and C. A. Conover. 1976. Chemical composition of good

quality tropical foliage plants. Proc. Fla. State Hort. Soc. 89:307-308. Poole, R. T. and C. A. Conover. 1982. Micronutrients in tropical foliage

production. Proc. Intern. Soc. Tropical Hort. 30:269-274.

Poole, R. T. and C. A. Conover. 1985a. A test of micronutrient sources.

Agr. Res. and Educ. Center - Apopka Res. Rpt. RH-85-4.

Poole, R. T. and C. A. Conover. 1985b. Boron and fluoride toxicity of foliage plants. Agr. Res. and Educ. Center - Apopka Res. Rpt. RH-85-

19.

Proc. Fla. State Hort. Soc. 104:303-308. 1991.

EFFECTS OF SEVERAL COMMERCIALLY AVAILABLE SOLIDIFYING AGENTS ON IN VITRO

GROWTH OF ALOCASIA BELLOTA 'ALICIA1

John L Griffis, Jr., Heike Wedekind,

and Sherman Johnson

Citrus Institute

Florida Southern College

111 Lake Hollingsworth Drive

Lakeland, FL 33801

Abstract. Cultures of Aloeasia be I lota 'Alicia' growing on hor

mone-free medium were divided and single shoots were

weighed and transferred to modified basic MS medium sup

plemented with 30 g/l sucrose, various organic compounds,

100 mg/l inositol, 0.5 mg/l IAA and 0.0, 1.0, or 10.0 mg/l 2iP.

Proc. Fla. State Hort. Soc. 104: 1991.

Media were solidified with one of seven different solidifying

agents including agar agar, Agaran™, Acaragel™, Caragar™,

carageenan and gellan gum (all from Research Organics) and

Gel rite™ (Kelco/Scott). Increases in fresh weights and num

bers of shoots and roots produced were measured every four

weeks for 120 days. Development of minimal callus was

noted with a few treatments, but vitrification was not noted

on any of the media combinations. The greatest increases in

both fresh weights and in shoot production occurred on media

supplemented with 0.5 mg/l IAA, 1.0 mg/l 2iP, and solidified

with either gellan gum or Gelrite™. Plants grown on media

solidified with agar, regardless of growth regulator concentra-

303

tion, showed the smallest fresh weight gains and produced

the fewest average numbers of new shoots.

Until about a decade ago, the only solidifying agents in

widespread usage in commercial plant tissue culture

laboratories were several different grades and brands of

agar (Pierik, 1987). Despite numerous claims from agar

manufacturers, few significant chemical differences have

ever been discovered between the various brands or types

of agar (Pierik, 1987), although different brands and dif

ferent concentrations of agar have been reported to have

significant effects on the growth and development of vari

ous plants in culture (Bornman and Vogelmann, 1984; De-

Bergh, 1983). It has also been reported that certain plants

may become vitrified when cultured on media solidified

with varying concentrations of agar (Bornman and Vogel

mann, 1984; DeBergh, 1983, Kevers et aL, 19894 Paques

and Boxus, 1987; Pasqueletto^a/., 1988a, & 1988b; Pierik,

1987; Turner and Singha, 1990; Zimmerman et aL, 1991;

Ziv, 1991). Increasing the agar concentration in the various

media often eliminated vitrification, but it also tended to

slow the rate of multiplication (DeBergh, 1983).

In 1983, Gelrite™, a gellan gum product, was intro

duced as an altertative gelling agent for tissue culture.

Both commercial plant tissue culture laboratories and

plant research laboratories around the world began to ex

periment with this new solidifying agent (Bornman and

Vogelmann, 1984; Ichi et aL, 1986; Kane et aL, 1990; Pas-

qualetto et aL, 1988a & 1988b; Pierik, 1987; Turner and

Singha, 1990; Zimmerman etaL, 1991). Several researchers

reported vitrification problems with Gelrite™ equal to or

even greater than those associated with agar (Bornman

and Vogelmann, 1984; Pasqualetto et aL, 1988a & 1988b;

Turner and Singha, 1990; Zimmerman et aL, 1991).

Numerous plants, including nandina, daylily, and gerbera

daisy (Griffis, unpublished data), Norway spruce

(Bornman and Vogelmann, 1984), apple (Pasqualetto et

aL, 1988a & 1988b), crabapple (Turner and Singha, 1990),

pear (Turner and Singha, 1990), geum (Turner and

Singha, 1990), and petunia (Zimmerman et aL, 1991) pro

duced vitreous tissues when cultured on various media sol

idified with Gelrite™. Increasing the Gelrite™ concentra

tion in the various media did not always eliminate vitrifica

tion (Bornman and Vogelmann, 1984). However, combi

nation of other agents with Gelrite™ sometimes corrected

the vitrification problem. A combination of Gelrite™ with

agar, both added to the medium at one-half of the man

ufacturer's recommended rate, eliminated the vitrification

of nandina shoots in culture (Griffis, unpublished data).

Similar results were obtained with petunia (Zimmerman et

aL, 1991). Others have also reported combining Gelrite™

and agar in culture media and obtaining satisfactory results

(Kane et aL, 1990). A commercial mixture of gellan gum

and agar, called Agaran™, is now available from Research

Organics of Cleveland, Ohio.

Other solidifying agents have also been suggested for

use in plant tissue cultures from time to time. Among those

suggested was carageenan (Ichi et aL, 1986), which is also

available from Research Organics as are a proprietary mix

ture of carageenan and agar, called Caragar™, and a blend

of carageenan and acacia gum, called Acaragel™. This last

blend always separated into distinct layers in the culture

vessels. Research Organics also markets their own brand

of agar, gellan gum, and agarose as "tissue culture medium

solidifiers."

This experiment was undertaken to evaluate and com

pare the effects of these various commercially available sol

idifying agents on a specific plant cultivar maintained for

several months in culture. The plant selected for this pro

ject was Alocasia bellota 'Alicia', an aroid of some minor

commercial value. However, other aroids, such as dieffen-

bachia, spathiphyllum, and syngonium, make up the vast

majority of plants propagated in commercial plant tissue

culture laboratories (Pierik, 1987) and data obtained from

this experiment with alocasia may be equally applicable to

other aroids. Solidifying agents selected for comparison

included Gelrite™ from Kelco/Scott and agar, carageenan,

Caragar™, Acaragel™, Agaran™, and gellan gum from Re

search Organics. Agarose was not trialed because it was at

least six times more expensive than any of the other sol

idifying agents and therefore, would not be cost-effective

for use by commercial laboratories even if it proved

superior to the others. Alocasia bellota 'Alicia' was main

tained in culture in a manner similar to that reported for

other aroids (Gomez et aL, 1989), except that side shoots

were not removed from the cultures during the course of

the four month experiment.

Materials and Methods

Multiplying cultures of Alocasia bellota 'Alicia' which had

been specially cultured on medium without plant hor

mones for thirty days were donated for this study by Dr.

Alvan Donnan and Plant Tech, Inc. of Apopka, Florida.

In the laminar flow environment, several single shoots

were removed from each culture. Each excised shoot was

carefully weighed and then cultured on one of twenty-one

different media. The various culture media were prepared

using modified MS basic salts (Sigma) supplemented with

30 g/1 sucrose, 2.0 mg/1 glycine, 0.5 mg/1 nicotinic acid, 0.5

mg/1 pyrodoxine HC1, 10.0 mg/1 thiamine HC1, 100 mg/1

myo-inositol, and 0.5 mg/1 indoleacetic acid (I A A). Cytoki-

nin levels were varied: media contained either 0.0, 1.0, or

10.0 mg/1 N6-(A2Isopentenyl) adenine (2iP). Media were solidified with one of seven different commercially avail

able solidifying agents including Agar Agar, Agaran™,

Acaragel™, Caragar™, Carageenan, and Gellan Gum (all

from Research Organics) or Gelrite™ (Kelco/Scott). Sol

idifying agents were used at the maximum rate suggested

by the supplier. These rates are sumamrized in Table 1

along with the coast of a kg of each solidifying agent

(purchased in 1990) and the coast of each solidifying agent/

liter of media. Single plantlets were cultured initially in 25

mm x 150 mm test tubes containing 10 ml of medium.

There were ten replicates of each treatment, 210 cultures

in all. After 30, 60, and 90 days in culture, plantlets were

removed from the various media, weighed and transferred

to fresh media. The number of additional shoots and roots

produced by each plantlet were also determined at each

transfer date. All transfers after the initial ones were made

to baby food jars (BFJ) each containing 25 ml of media.

After 120 days in culture, final measurements were taken

and cultures were terminated. During the course of the

experiment, cultures were maintained at 24 ± 1°C under

a 16-hour photoperiod per day provided by cool-white

fluorescent tubes at 60 (xmol-s-1 nrr2 as measured at the cul-

304 Proc. Fla. State Hort. Soc. 104: 1991.

Table 1. Solidifying Agent Cost Comparison.

Solidifying agent

Acaragel™ (ROZ

AgarAgar(RO)

Agaran™ (RO)

Caragar™ (RO)

Carageenan (RO)

Gellan Gum (RO)

Gelrite™ (Kelco/Scotty)

Gelrite™ (Kelco/Scott)

Cost/kg

$54.00

$41.00

$54.00

$54.00

$20.00

$83.00

$90.00

$90.00

Recommended

Rate

10g/l

8g/l

5g/l

6g/l

10g/l

3g/l 3g/l 2g/l

liters of

media/kg

100

125

200

166

100

333

333

500

actual cost

/liter

$0.54/1

$0.33/1

$0.27/1

$0,325/1

$0.20/1

$0.25/1

$0.27/1

$0.18/1

Note: Culture vessels (baby food jars) hold approximately 25 ml of media,

so 1 liter of media will fill 40 vessels. From the above figures, it can be

determined that the cost of solidifying agent varies from about 0.5tf to

1.5tf per vessel.

Z(RO); Research Organics of Cleveland, Ohio.

y( Kelco/Scott); Kelco is a division of Merck and Co., Rahway, NJ.

ture level. Data were analyzed using the ANOVA proce

dure.

Results and Discussion

Roots

There were significant differences among the 21 treat

ments in average numbers of roots produced after one,

two, three, and four 30-day culture periods, hereafter re-

fered to as 30, 60, 90, and 120 days in culture (Table 2).

As might be expected, the cultures maintained on MS

medium supplemented with 0.5 mg/1 IAA, but no cytoki-

Table 2. Comparative production of roots in vitro by Alocasia bellota 'Alicia'.

Solidifying agent/

cytokinin level

Acaragel™/0 mg/1 2iP



Agar Agar/ 0 mg/1 2iP

Agar Agar/1 mg/1 2iP


Agaran™/0 mg/1 2iP


Agaran™ 10/mg/12iP

Caragar™/ 0 mg/1 2iP

Caragar™/1 mg/1 2iP


Carageenan/ 0 mg/1 2iP

Carageenan/1 mg/1 2iP


Gellan Gum/ 0 mg/1 2iP

Gellan Gum/1 mg/1 2iP


Gelrite™/0 mg/12iP Gelrke™/1 mg/1 2iP

Gelrite™/10 mg/1 2iP

ANOVA: Pr>Fat.O5

30 Days

1.80z

0.60

0

0.90

0.20

0

0.90

0.60

0

1.20

0.40

0

1.70

0.60

0

2.20

1.50

0

0.90

1.80

0.10

0.000

60 Days

1.90

0.70

1.33

2.33

0.50

0

2.20

0.30

0.30

1.22

0.43

0.60

2.50

0.50

0

2.11

2.11

2.78

7.89

1.33

1.33

0.011

90 Days

2.78

0.50

1.22

1.00

0.22

0.22

2.25

0.80

1.70

1.88

0.43

0.67

4.11

2.75

0.25

2.50

10.17

5.11

14.88

9.25

4.63

0.000

120 Days

2.43

2.88

1.56

2.14

2.00

1.00

2.50

2.67

3.25

1.86

1.17

1.25

4.89

8.00

0.50

3.13

17.67

6.71

2.43

18.57

7.25

0.000

zValues in each column represent the average increase in number of

roots produced per treatment. All values are cumulative from day zero

up to the evaluation date listed at the top to each column. The initial

explants had no roots and there were initially ten replicates of each treat

ment. A decrease in the average number of roots produced by any treat

ment over time may have occurred because one or more cultures of that

treatment were lost to contamination during the experiment.

Proc. Fla. State HorL Soc. 104: 1991.

nin, regardless of solidifying agent, demonstrated the

greatest initial increases in average numbers of roots pro

duced. Cultures maintained on medium supplemented

with 0.5 mg/1 IAA and 10 mg//l 2iP, regardless of solidify

ing agent, produced the lowest average numbers of roots

at each evaluation. Cultures maintained on medium sup

plemented with 0.5 mg/1 IAA and 1 mg/1 2iP and solidified

with either gellan gum or Gelrite™ had produced the

greatest average numbers of roots at the end of 120 days.

Moreover, these last two particular combinations of growth

regulators and solidifying agents also produced the great

est averge number of shoots in 120 days and many of the

new shoots also had begun to produce roots at the time the

experiment was terminated.

Fresh Weight

There were also significant differences among the 21

treatments in percentage fresh weight gain after 30, 60,

90, and 120 days in culture (Table 3). Not surprisingly, the

cultures maintained on MS medium supplemented with

0.5 mg/1 IAA, but no cytokinin, regardless of solidifying

agent, produced the smallest increases in average percent

fresh weight gain at each evaluation. Numerous cultures

actually demonstrated decreases in fresh weight after three

or four transfers onto media without cytokinin. On the

other hand, cultures maintained on medium supple

mented with 0.5 mg/1 IAA and 10 mg/1 2iP, regardless of

solidifying agent, produced substantial increases in aver

age percent fresh weight gain at each evaluation. Notably,

however, cultures maintained on medium supplemented

with 0.5 mg/1 IAA and only 1 mg/1 2iP and solidified with

either gellan gum or Gelrite™ produced the greatest aver

age increases in percent fresh weight gain at every evalua tion.

Shoots

There were also significant differences among the 21

treatments in average numbers of shoots produced after

30, 60, 90, and 120 days in culture. The ANOVA pro

duced a Pr>F (.05) of 0.000 at all four evaluations. The

cultures maintained on MS medium supplemented with 0.5 mg/1 IAA, but no cytokinin, regardless of solidifying

agent, produced the fewest average number of shoots at

each evaluation. Numerous explants failed to produce a

single new shoot after 120 days in culture when cytokinin

was left out of the culture medium (Fig. 1). When 2iP was

included in the culture medium at either 1 mg/1 or 10 mg/1,

substantial average numbers of new shoots were produced

in culture, regardless of the solidifying agent. Average shoot production was almost identical at either 2iP level

after 60 days and 90 days. Only after 120 days in culture was there a large difference in the average number of

shoots produced per culture between the two cytokinin treatments.

Cultures maintained on MS medium solidified with

agar, regardless of cytokinin level, produced the smallest

increase in average numbers of new shoots at each evalua tion (Fig. 2). Cultures maintained on medium solidified with either gellan gum or Gelrite™, regardless of cytokinin

level, produced the greatest increase in average numbers of new shoots at each evaluation.

305

Table 3. Comparative percent fresh weight gain in vitro of Alocasia bellota 'Alicia'.

Solidifying agent/

cytokinin level 30 Days 60 Days 90 Days 120 Days

Acaragel™/0mg/12iP


Acaragel™/10mg/12iP




Agaran™/0mg/l2iP


Agaran™ 10/mg/12iP

Caragar™/0mg/12iP


Caragar™/10mg/12iP

Carageenan/ 0 mg/1 2iP

Carageenan/1 mg/12iP


Gellan Gum/ 0 mg/1 2iP


Gellan Gum/10 mg/12iP

Gelrite™/0mg/12iP

Gelrite™/1 mg/1 2iP

Gelrite™/10mg/12iP

194.89%z

102.82%

148.05%

72.55%

60.86%

89.75%

123.27%

89.29%

134.35%

162.82%

131.62%

161.67%

147.96%

178.36%

129.04%

149.96%

316.72%

289.72%

181.92%

404.84%

249.41%

343.11%

375.78%

683.05%

24.55%

167.67%

455.40%

383.74%

342.15%

1323.34%

420.15%

453.00%

756.56%

523.49%

852.50%

867.19%

414.50%

1997.92%

1348.96%

492.03%

2673.68%

1244.94%

598.72%

536.20%

1896.55%

-97.00%

334.32%

1115.50%

718.94%

688.16%

1837.51%

210.22%

553.77%

2170.53%

855.38%

963.62%

1707.67%

554.36%

5383.77%

3859.47%

1423.30%

8190.36%

4701.81%

117.15%

1575.04%

5238.19%

-223.16%

758.44%

3095.45%

1153.33%

1582.01%

5411.30%

-126.76%

1094.42%

6636.13%

1018.08%

3396.14%

5297.02%

718.18%

12940.17%

11396.36%

1792.88%

16010.13%

9010.94%

ANOVA: Pr>F at .05 0.000 0.011 0.000 0.000

zValues in each column represent the average increase fresh weight gain per treatment during the culture period. The variation in the initial fresh

weights of the explants has been taken into account. There were initially ten replicates of each treatment.

When the combined effects of cytokinin level and so

lidifying agent were examined more thoroughly, several

trends were discovered. Cultures maintained on MS

medium supplemented with 1 mg/1 2iP and solidified with

either gellan gum or Gelrite™ produced the greatest in

creases in average numbers of shoots at each evaluation

when compared with other solidifying agents trialed (Fig.

3). However, cultures maintained on medium supple

mented with 10 mg/1 2iP produced very nearly the same

increases in average numbers of shoots at each evaluation

no matter which solidifying agent was used (Fig. 4). Even

at the higher 2iP level, agar remained the poorest per

former of all solidifying agents trialed when evaluted for

increase in average numbers of shoots produced at each

evaluation.

A comparison of cultures maintained on MS medium

supplemented witn various levels of 2iP and solidified with

either gellan gum, Gelrite™, or agar produced some in

teresting comparisons (Fig. 5). There was little difference

Gelrlte ||j||||)j^ | |

Gellan Gum IHBiMHHlHsIS'

Carageenan wsSBmEggP^'

Caragar teaftjmfflf*[^

1 j ) 1 Agaran |H|

Agar Wfflf

Acaragel &&£&&

■n

1

(ff{

H j

0 120 days

■ 90 days

H 60 days

■ 30 days

10 12 14

Average Increase in Numbers of Shoots per Culture

Fig. 2. Effects of Solidifying Agent on Average Numbers of Shoots

Produced In Vitro by Alocasia bellota 'Alicia'.

IOmg/1 21P

I mg/1 21P

0mg/l 21P

120 days

90 days

60 days

30 days

Gelrite 1 mg/1 2iP

Gellan Gum 1 mg/1 2iP

Carageenan 1 mg/1 2iP

Caragar 1mg/l 2iP

Agaran 1 mg/1 2iP

Agar 1 mg/1 21P

Acaragel 1 mg/1 2iP

120 days

90 days

60 days

30 days

0 4 8 12 16


0 3 6 9 12 15 18 21


Figure 3. Combined Effects of Both Solidifying Agent and Cytokinin

Fig. 1. Effect of 2iP Levels on Average Numbers of Shoots Produced Level (2iP 1 mg/1) on Average Numbers of Shoots Produced In Vitro by In Vitro by Alocasia bellota 'Alicia'. Alocasia bellota 'Alicia'.

306 Proc. Fla. State Hort. Soc. 104: 1991.

120 days

90 days

60 days

30 days

Gelrite IOmg/1 21P

GellanGum IOmg/1 2iP

Carageenan 10mg/l 2iP

Caragar IOmg/1 2iP

Agaran IOmg/1 2iP

Agar IOmg/1 2iP

Acaragel, 10mg/l 2iP

0 3 6 9 12 15 18 21


Fig. 4. Combined Effect of Both Solidifying Agent and Cytokinin

Level (2iP 10 mg/1) on Average Numbers of Shoots Produced In Vitro by

Alocasia bellota 'Alicia'.

Gellan Gum 10 mg/l 2iP

Gellan Gum I mg/l 2iP

Gellan Gum 0mg/l 2IP

Agaran 10mg/l 2iP

Agaran I mg/l 21P

Agaran 0mg/l 2IP

Agar IOmg/1 2IP

Agar lmg/l 2iP

Agar 0mg/l 2IP

120 days

90 days

60 days

30 days

0 3 6 9 12 15 18 21


Fig. 6. Combined Effect of Both Selected Solidifying Agents (Gellan

Gum, Agar, and Agaran™) and Various Cytokinin Levels on Average

Numbers of Shoots Produced In Vitro by Alocasia bellota 'Alicia'.

between gellan gum and Gelrite™ in average numbers of

shoots produced at any evaluation. Agar was clearly in

ferior to the other two gelling agents when any level of 2iP

trialed was added to the culture medium, but when cytoki

nin was excluded from the medium, none of the three

solidifying agents differed significantly in the average

numbers of shoots produced at each evaluation. The addi

tion of the higher 2iP level (10 mg/l) to the culture medium

produced a four-fold increase in the average numbers of

shoots produced on the medium solidified with agar after

120 days when compared to medium supplemented with

only 1 mg/l 2iP. However, the inclusion of 10 mg/l 2iP did

not produce any increase in average numbers of shoots

produced on medium solidified with either gellan gum or

Gelrite™ after 120 days when compared to medium sup

plemented with only 1 mg/l 2iP. In fact, there was no dif

ference at all in average numbers of shoots produced be

tween cultures maintained on MS medium supplemented

with either 1 mg/l 2iP or 10 mg/l 2iP and solidified with

gellan gum after 120 days in culture. There was an actual

decrease in average numbers of shoots produced at all

evaluations when the higher level of 2iP was added to

medium solidified with Gelrite™ when compared to

medium supplemented with the lower level of 2iP and sol

idified with Gelrite™ (Fig. 5).

A comparison of cultures maintained on MS medium

supplemented with various levels of 2iP and solidified with

either agar, gellan gum, or Agaran™ also produced some

interesting comparisons (Fig. 6). Agaran™ is a proprietary

Gelrite 10 mg/l

Gelrite lmg/l

Gelrite 0mg/1

Gellan Gum 10mg/l

GellanGum lmg/l

GellanGum 0mg/l

Agar 10mg/l

Agar lmg/l

Agar 0mg/l

120da>s

90 days

60 days

30 days

0 3 6 9 12 15 18 21


Fig. 5. Combined Effect of Both Selected Solidifying Agents (Gel

rite™ , GeWan Gum, and Agar) and Various Cytokinin Levels on Average

Numbers of Shoots Produced In Vitro by Alocasia bellota 'Alicia'.

Proc. Fla. State Hort. Soc. 104: 1991.

combination of agar and gellan gum and its performance

as a solidifying agent was intermediate between the other

two agents when evaluated for average numbers of shoots

produced at all evaluations. Among these nine variations,

cultures maintained on medium supplemented with 1 mg/l

2iP and solidified with gellan gum produced the greatest

increases in average numbers of shoot at all evaluations.

Production Comparisons

Examinations of the various experimental data sets pre

sented above have yielded several important production

comparisons that may be of some commercial interest.

Some of them are listed below.

1) Without cytokinin in the media, there was little dif

ference between the solidifying agents; none could pro

duce a satisfactory increase in average numbers of shoots

of Alocasia bellota 'Alicia' per culture even after four 30-day

culture periods.

2) With the addition of 1 mg/l 2iP to the media, only

gellan gum or Gelrite™ produced a commercially accept

able increase in average numbers of shoots (>3) of Alocasia

bellota 'Alicia' per culture in the initial 30-day culture

period.

3) With the addition of 1 mg/l 2iP to the media, all

other solidifying agents trialed required either two or three

30-day culture periods to produce a reasonable increase in

average numbers of shoots (>3) per culture except agar,

which required four 30-day culture periods to produce a

reasonable increase in average numbers of shoots (>3) per

culture.

4) With the addition of 10 mg/l 2iP to the media, there

was initially little difference between solidifying agents

trialed; none could produce an increase in average num

bers of shoots per culture during the initial 30-day culture

period as great as that achieved in certain Alocasia bellota

'Alicia' cultures grown on media supplemented with only 1 mg/l.

5) Two or three 30-day culture periods of growth on

media supplemented with 10 mg/l 2iP did not substantially

increase the average number of shoots produced per cul

ture compared to the number of shoots produced on

media supplemented with only 1 mg/l 2iP, except in the

case of Agaran™, where average number of shoots pro

duced was doubled on media supplemented with the

higher level of 2iP.

307

6) Four 30-day culture periods on media supplemented

with 10 mg/1 2iP substantially increased the average num

bers of shoots produced per culture compared to the aver

age number of shoots produced on media supplemented

with 1 mg/1 2iP, except in the cases of Gelrite™ and gellan

gum, where shoot production was generally lower on

media supplemented with the higher level of 2iP.

Conclusions

1) Vitrification was a primary concern when this exper

iment was initiated. However, neither the addition of 1

mg/1 nor 10 mg/1 of 2iP to the culture medium caused

vitrification of any of the Alocasia bellota 'Alicia' plantlets

trialed, even after four 30-day culture periods. Solidifying

agents did not seem to have any effect on vitrification in

this experiment.

2) Callus was not a significant problem in any of the

Alocasia bellota 'Alicia' cultures, although small amounts of

callus were formed in several of the cultures after four

30-day culture periods on media supplemented with 10

mg/1 2iP. Solidifying agents did not seem to have any effect

on callus production in this experiment.

3) The most cost-effective combinations of cytokinin

and solidifying agent were 1 mg/1 2iP and either gellan

gum or Gelrite™ when cultures were evaluated for either

average shoot production or average fresh weight increase.

Cost of 2iP and either of these two solidifying agents would

be about 1# per culture vessel (BFJ). Both of these cytoki

nin and solidifying agent combinations yielded an average

of slightly more than nine shoots per culture after three

30-day culture periods and an average of more than 17

shoots per culture after four 30-day culture periods. All

other cytokinin and solidifying agent combinations trialed

in this experiment either cost more, produced fewer shoots

over time, or both.

Bibliography

1. Bornman, C. H. and T. C. Vogelmann. 1984. Effect of rigidity of gel

medium on benzyladenine-induced adventitious bud formation and

vitrification in vitro in Picea abies. Physiol. Plant. 61:505-512.

2. DeBergh, P. C. 1983. Effects of agar brand and concentration of the

tissue culture medium. Physiol. Plant. 59:270-276.

3. Gomez, L., M. Monge, R. Valverde, O. Arias, and T. Thorpe. 1989.

Micropropagacion de tres araceas comestibles libres de virus. Tur-

rialba 39:155-161.

4. Ichi, T., T. Koda, I. Asai, A. Hatanaka, and J. Sekiya. 1986. Effects

of gelling agents upon in vitro culture of plant tissues. Agric. Biol.

Chem. 50:2397-2399.

5. Kane, M. E., N. L. Philman, and T. M. Lee. 1990. Evaluation of the

Cultusak™ in vitro plant culture system. Proc. Fla. State Hort. Soc.

103:182-186.

6. Kevers, C, M. C. Coumans, M.-F. Coumans-Gilles, and Th. Gaspar.

1984. Physiological and biochemical events leading to vitrification of

plants cultured in vitro. Physiol. Plant. 61:69-74.

7. Paques, M. and Ph. Boxus. 1987. "Vitrification": review of literature.

Acta Horticulturae 212:155-166.

8. Pasqualetto, P.-L., W. P. Wergin, and R. H. Zimmerman. 1988a.

Changes in structure and elemental composition of vitrified leaves of

'Gala' apple in vitro. Acta Horticulturae 227:352-357.

9. Pasqualetto, P.-L., R. H. Zimmerman, and I. Fordham. 1988b. The

influence of cation and gelling agent concentrations on vitrification

of apple cultivars in vitro. Plant Cell, Tiss. and Org. Cult. 14:31-40.

10. Pierik, R. L. M. 1987. In vitro culture of higher plants. Martinus

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11. Turner, S. R. and S. Singha. 1990. Vitrification of crabapple, pear,

and geum on gellan gum-solidified culture medium. HortScience

25:1648-1650.

12. Zimmerman, T. W., S. M. D. Rogers, and B. G. Cobb. 1991. Control

ling vitrifiation of petunia in vitro. In Vitro Cell. Dev. Biol. 27P: 165-

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Proc. Fla. State Hort. Soc. 104:308-310. 1991.

SUBSTRATE INFLUENCE ON COMPOST EFFICACY AS A NURSERY GROWING MEDIUM

George E. Fitzpatrick and Stephen D. Verkade

University of Florida

Fort Lauderdale Research and Education Center

3205 College Avenue

Fort Lauderdale, Florida 33314

Additional index words, waste management, municipal solid

waste, yard trash, sewage sludge.

Abstract. Composts made from municipal solid waste (MSW),

used disposable baby diapers (DIAP), horse stable waste (StW),

yard trash (YT), and a co-compost made from dewatered sew-

Florida Agricultural Experiment Stations Journal Series No. N-00581.

Portions of this research were supported by grants from Agripost, Inc.,

the Florida State Center for Hazardous and Solid Waste Management,

and the Horticultural Research Institute. We thank William R. Farrell of

the Broward County Pubic Works Department for providing the yard

trash compost and the sludge-yard trash co-compost; Ted Malloy, formerly

of the Pompano Harness Track, for providing the stable waste compost;

James McNelly, formerly of Recomp, Inc., for providing the disposable

diaper compost and John Nowell, formerly of Agripost, Inc., for providing

the municipal solid waste compost. We gratefully acknowledge the techni

cal assistance provided by Jane E. Slane and Richard R. Weidman.

308

age sludge and yard trash (1:3, by volume) (S-YT) were used

both as stand-alone growing media and as ingredients in

media blends for viburnum, Viburnum suspensum. Plants

were grown for 5 1/2 months in 25 cm. diameter containers

with a commercially available growing medium (4 sedge peat:

5 pine bark: 1 sand, by volume) as a control. When used as

a stand-alone medium, the S-YT produced plants that were

significantly taller and had significantly greater biomass than

plants grown in any of the other media, including the control

medium. The S-YT, when blended with pine bark and sand,

produced larger plants than did the blends using MSW and

YT, but the differences were smaller. Medium pH, total conduc

tivity, % pore space and % water holding capacity did not

seem to be as strongly related to efficacy as the relative level

of medium subsidence during production. The best performing

medium, S-YT, compacted the least during the production

period and the media that performed the poorest had the

greatest levels of compaction during the production period.

The issues of urban waste management on one hand

and the need of commercial horticultural production

facilities to have continued supplies of plant growing media

Proc. Fla. State Hdrt. Soc. 104: 1991.

EFFECTS OF SEVERAL COMMERCIALLY AVAILABLE SOLIDIFYING ...

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