Field Transplant Survival of Amelanchier Liners Produced by Tissue Culture

Daniel K. Struve and R. Daniel Lineberger

ABSTRACT

Tissue cultured Amelanchier laevis liners were transplanted to the field with almost complete survival. Plants transplanted from Cone-Trainers were slightly shorter, but had more lateral breaks 1 year after planting. A period of outdoor acclimation under 60% shade for as little as 5 days reduced transplant shock as indicated by incremental growth at 1, 2, or 3 months, but plant height and number of lateral breaks after 1 year were not affected by outdoor acclimation. Tissue cultured liners could be outplanted as late as early October and still overwinter without damage.

INTRODUCTION

Propagation of nursery crops through tissue culture is gaining increasing acceptance from the nursery industry. Micropropagation systems from initial explanting to acclimation of microcuttings have been developed for many species [see Lineberger (3) for an example of such a system]. Physiological and morphological changes which occur in micropropagated plants during acclimation from aseptic conditions to the greenhouse environment have focused on two areas. The structure and amount of epicuticular was on the leaves increases upon exposure of microcuttings to lowered relative humidity, a change deemed necessary to lower rates of water loss from the delicate microcuttings (5). Concomitantly, the stomata increase in rate of response to water stress after 5 days of acclimation. This change is believed to be significant to the survival of tissue cultured woody plants (1).

Relatively little research has been conducted to determine factors necessary to acclimation of tissue cultured liners to the field environment. Smith (4) recommends a minimum size of 10-15 cm top growth and an 8-10 cm root ball for field transplanting of tissue cultured liners. Dunstan (2) reported that tissue cultured apple rootstocks transplanted to the field directly from the greenhouse in early spring were susceptible to frost damage and were defoliated but regrew after about 2 months of quiescence. DunstanÕs transplanting procedure for summer transplanting now involves preconditioning plants under 50% shade to achieve a Ņpartially exposed environment into which plants are placed before full exposure.Ó Losses of tissue cultured liners were no greater than those for seedlings using this procedure.

Relatively little research has been conducted to determine factors necessary to acclimation of tissue cultured liners to the field environment. Smith (4) recommends a minimum size of 10-15 cm top growth and an 9-10 cm root ball for field transplanting of tissue cultured liners. Dunstan (2) reported that tissue cultured apple rootstocks transplanted to the field directly from the greenhouse in early spring were susceptible to frost damage and were defoliated but regrew after about 2 months of quiescence. DunstanÕs transplanting procedure for summer transplanting now involves preconditioning plants under 50% shade to achieve a Ņpartially exposed environment into which plants are placed before full exposure.Ó Losses of tissue cultured liners were no greater than those for seedlings using this procedure.

This research was undertaken to determine the effect of certain factors (container type, planting date, days of outdoor acclimation) on the transplant survival of tissue cultured plants of Amelanchier laevis.

MATERIALS AND METHODS

Microcuttings of Amelanchier laevis were produced in vitro according to the method described for ŌHally JolivetteÕ cherry (3). The growth regulator concentration in the shoot proliferation stage was 0.1 mg/1 naphthaleneacetic acid and 2.5 mg/1 benzladenine. These microcuttings were rooted in plastic-covered foil containers containing a moistened peatmoss-perlite medium (1:1 by volume). Rooting was done in the tissue culture laboratory where the temperature was 24-27”C and lighting was provided for 16 hr per day at 40µ Einsteins/M2/sec by cool white fluorescent lamps.

Transplanted microcuttings were immediately placed on a shaded intermittent mist bench for 3 days (6 sec most every 6 min; light intensity, 360µ Einsteins/M2/sec). After an additional 7 to 10 days of acclimation on a shaded greenhouse bench (light intensity, 270µ Einsteins/m2/sec), plants were grown under standard greenhouse cultural conditions including fertilization at every watering with 200 ppm of 20-20-20 soluble fertilizer. Plants which received an outdoor acclimation treatment were held for various time intervals under 60% shade (light intensity, 810µ Einsteins/m2/sec) prior to planting.

Field transplanting studies were conducted on the Ohio State University Dept. of HorticultureÕs Lane Avenue Farm. The plot used was a Brookston silty clay loam with a previous record of heavy fertilization. The plot was irrigated as needed.

In experiment 1, plantlets approximately 15 cm tall were acclimated for 0, 5, 10, or 15 days under 60% shade outdoors. Two types of containers were used, 5.7 x 5.7 cm peatmoss pots or 4 x 20 cm Cone-Tainers (Ray Leach Cone-Tainer Nursery, Canby, OR 97013). A 1:1 (by volume) peatmoss-perlite medium was used. Field planting was done June 28, 1981. A randomized complete block design with six six-plant replications was used. Experiment 2 was a replication of experiment 1 except only peatmoss pots were used. Field planting was done August 6, 1981. A randomized complete block design was used with six six-plant replications. Experiment 3 was conducted to determine how late in the year plants could be transplanted without suffering significant winter damage. At approximately 3-day intervals beginning Sept. 6 and ending Oct. 4, 1981, ten 15 cm tall plants growing in peatmoss pots were transplanted directly from the greenhouse to the field.

In all experiments, plant height, number of leaves, and number of basal lateral branches were recorded. Plants in experiments 1 and 2 were measured monthly until Oct. 4, 1981, and again on June 24, 1982, after the first growth flush ceased elongation. Winter survival was assessed in May and June 1982.

All data were subjected to analysis of variance. Mean separation by DuncanÕs multiple range test was conducted only for effects significant at the 0.05 level.

RESULTS AND DISCUSSION

Tissue cultured plants of Amelanchier laevis survived transplanting in excellent condition. When results are compared by container type across all other treatments, transplant losses were negligible in either container (Table 1).

Table 1. Comparison of the Transplant Survival, Shoot Growth, and Basal Branching of Tissue Cultured Amelanchier laevis Transplanted to the Field from Peatmoss Pots or Cone-Tainers.

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								Number of
Container		Percentage	Increase in		Basal Lateral
   Type		Survival*	Shoot Length (cm)**	Branches***
						

Cone-Tainer		99%		59.3a****		2.9a

Peatmoss Pot		98%		64.4b			1.8b
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Planting date: June 28, 1981. *Percentage of 144 plants which survived; survival was the same 3 or 12 months after transplanting. **Increase in shoot length from June 28, 1981, to June 24, 1982. ***Average number of basal lateral branches observed 4 months after field planting. ****Means within columns followed by different letters are significantly different at the 0.05 level by Duncan's multiple range test.

Shoot growth was slightly higher when plants were grown in peat pots, but those transplanted from Cone-Tainers had a higher number of basal lateral branches at the end of the first growing season (Table 1).

Plants exposed to a period of outdoor acclimation under shade prior to field planting appeared to have less transplant shock. Plants acclimated for 5 to 10 days exhibited the greatest incremental increase in height after 1 month (Table 2).

Table 2. Height Increase and Number of Basal Lateral Shoots of Tissue Cultured Amelanchier laevis Transplanted to the Field Following Various Time Intervals of Holding Outdoors Under 60% Shade Prior to Planting.

										
Days 				Height	Basal
Outdoor	Initial		Incremental Height Increase (cm)	at 12	Lateral
Accl		Height	1 mo	2 mo	3 mo	12 momonths (cm)	Branches
								

0		24.6	5.5a*	13.4b	14.8a	59.3ab	83.9	2.3a
5		19.7	7.5c	16.9c	18.8b	62.7b	82.4	2.2a
10		19.6	7.1c	14.4b	16.7ab	66.1b	85.7	2.6a
15		18.5	6.0b	11.7a	14.3a	59.0a	77.5	2.6a

Planting date: June 28, 1991. *Means within columns followed by the same letter are not significantly different at the 0.05 level by DuncanÕs mean separation. During the 1, 2, and 3 month measurement periods, those plants acclimated for 5 days grew more than those acclimated for 10 days, which in turn grew more than those acclimated for 15 days (differences were not all significant, but the trend was evident). By the end of 12 months, the growth rates of all plants tended to one value despite pretransplant acclimation treatment (Table 2). Additionally, the acclimation treatments did not affect the number of basal lateral shoots observed 4 months after planting (Table 2).

When the experiment was repeated with an August rather than a June planting date (experiment 2) and using only plants grown in peat pots, a slight benefit to prior acclimation was observed. Increasing time outdoors under shade increased both growth in height and the number of basal lateral shoots observed after 3 months (Table 3).

The experiment was terminated before the long term effects of acclimation on August transplanting were determined. Survival was 100%. A final study was conducted to determine how late in the season transplanting could be done without affecting overwinter survival (experiment 3). These plants were transplanted to the field directly from the greenhouse as late as Oct. 4, 1982. When survival data were taken on May 12, 1983, it was determined that all transplants had survived the winter and begun active shoot growth.

The number of leaves per plant at transplanting had no effect on transplant or winter survival or on height growth in any of these experiments. It was common for the lower third of the leaves to abscise within 2 weeks of transplanting.

These studies have demonstrated excellent field transplant survival of tissue cultured Amelanchier laevis. Survival levels of nearly 100% can be achieved: 1) without outdoor acclimation under shade, 2) when plants are grown either in peatmoss pots or Cone-Tainers, and 3) at planting dates varying from late June to early October. A period of outdoor acclimation appears to lessen transplant shock of either June or August transplanting, but the long term (12 month) benefit to this procedure with regard to shoot growth or basal branching appears negligible.

These results should not be generalized to plants which are succulent at the time of transplanting, however. Even though the Amelanchier liners used in this study were greenhouse grown, they did have sturdy, woody stems at transplanting.

LITERATURE CITED

1. Brainerd, K.E. and L.H. Fuchigami. 1981. Acclimatization of aseptically cultured apple plants to low relative humidity. J. Amer. Soc. Hort. Sci. 106:515-518.

2. Dunstan, D.I. 1981. Transplantation and post-transplantation of micropropagated tree fruit rootstocks. Proc. Int. Plant Prop. Soc. 31:39-45.

3. Lineberger, R.D. 1983. Shoot proliferation, rooting, and transplant survival of tissue-cultured ŌHally JolivetteÕ cherry. HortScience 18:182-185.

4. Smith, W.A. 1981. The aftermath of the test tube in tissue culture. Proc. Int. Plant Prop. Soc. 31:47-49.

5. Sutter, E. and R.W. Langhans. 1979. Epicuticular wax formation on carnation plantlets regenerated from shoot tip culture. J. Amer. Soc. Hort. Sci. 104:493-496.

Table 1. Comparison of the Transplant Survival, Shoot Growth, and Basal Branching of Tissue Cultured Amelanchier laevis Transplanted to the Field from Peatmoss Pots or Cone-Tainers. Number of Container Percentage Increase in Basal Lateral Type Survival* Shoot Length (cm)  Branchesą Cone-Tainer 99% 59.3a** 2.9a Peatmoss Pot 98% 64.4b 1.8b Planting date: June 28, 1981. *Percentage of 144 plants which survived; survival was the same 3 or 12 months after transplanting.  Increase in shoot length from June 28, 1981, to June 24, 1982. ąAverage number of basal lateral branches observed 4 months after field planting. **Means within columns followed by different letters are significantly different at the 0.05 level by DuncanÕs multiple range test. Table 2. Height Increase and Number of Basal Lateral Shoots of Tissue Cultured Amelanchier laevis Transplanted to the Field Following Various Time Intervals of Holding Outdoors Under 60% Shade Prior to Planting. Number Days of Height of Basal Outdoor Initial Incremental Height Increase (cm) at 12 Lateral Acclimation Height 1 month 2 months 3 months 12 months months (cm) Branches 0 24.6 5.5a* 13.4b 14.8a 59.3ab 83.9 2.3a 5 19.7 7.5c 16.9c 18.8b 62.7b 82.4 2.2a 10 19.6 7.1c 14.4b 16.7ab 66.1b 85.7 2.6a 15 18.5 6.0b 11.7a 14.3a 59.0a 77.5 2.6a Planting date: June 28, 1991. *Means within columns followed by the same letter are not significantly different at the 0.05 level by DuncanÕs mean separation. Table 3. Height Increase and Basal Branching of Tissue Cultured Amelanchier laevis Subjected to Various Periods of Outdoor Acclimation Prior to Field Planting. Height Days of Increase (cm) Number of Outdoor Initial After Basal Acclimation Height (cm) 2 Months Lateral Shoots 0 11.7 0.0b* 1.5a 5 10.4 0.2b 1.4a 10 9.7 2.6a 1.9b 15 10.1 4.0a 2.1b Planting date: August 6, 1981. *Means within columns followed by the same letter are not significantly different at the 0.05 level by DuncanÕs mean separation. 1Partial support for this project was provided by grants from the Horticultural Research Institute and the Western Region of the International Plant Propagators Society. The authors thank Joe Takayama for his excellent technical support. 2Assistant Professor and Associate Professor, Department of Horticulture.