Research Protocol

Acclimatization of Tissue Culture Plants

Sweetpotato: Acclimatization of Tissue Culture Plants

Authors: Srijana Shrestha, Jessica Weaver, Carol Miles
Affiliation: Washington State University Northwestern Washington Research and Extension Center

_{https://vegetables.wsu.edu/sweetpotato/}_{November 2025}

Introduction

Sweetpotato production is limited in northern regions due to the perceived barriers of a short growing season and relatively cool summer temperatures. Yet recent studies have shown sweetpotatoes grown with plastic mulch in northern regions produced greater yield than the national average (11 tons/acre) due to the longer photoperiod during the growing season. Thus, in areas such as Mount Vernon, WA where day length is 16 h on average in June and July, rapid growth during establishment of healthy transplants likely contributes to higher yields. In a preliminary study at Northwestern Washington Research and Extension Center, Mount Vernon, WA, the yield of some sweetpotato accessions ranged from 14-26 tons/acre in 2020. The keys to successful production are suitable cultivars, healthy slips, planting into warm soil, appropriate plant spacing, and maintaining warm soil throughout the growing season.

Hardening off tissue culture plants

Sweetpotatoes are propagated vegetatively on a large commercial scale by slips or vine-cuttings. However, disease-causing pathogens and pests can be easily transferred from one generation to the next by vegetative propagation. Moreover, sweetpotatoes are greatly susceptible to mutations that pass through generations. When the same genetic stock is used for several years, off-type planting materials can be propagated and cause reduction in yield. Therefore, it is advised to replace the seed stock with new certified disease-free stock for slip production every three years. Tissue culture plants ensure disease-free planting materials but require greater attention to prepare for transplanting to the bed. Plant tissue culture is carried out under sterile and controlled conditions of light, temperature, and humidity. Thus, a transitional period of growth and acclimatization is required before transplanting.

Supplies

70% ethanol — to sanitize surfaces, tools, and hands
Sterlized latex gloves — to prevent contamination
4-inch pots — to hold plants
Potting media — to cover plant roots
Forceps — to remove plants from tissue culture tubes
Fungicide — to avoid fungal infection
Humidity box — to maintain optimal growing conditions

Experimental set up

Before planting tissues culture plants in the field, they must be grown in a greenhouse and acclimate to the outdoor environment in a hoop house.

Steps to acclimatize tissue culture plants

Receive the packing box in a clean and disinfected area to reduce the possibility of contamination. Wipe the counter surface with 70% ethanol where the box will be placed and also wipe the outside of the box before opening.
Remove the culture tubes from the packing box while wearing sterile latex gloves.
Place the tubes at room temperature 22-24 °C with 1000 lux illumination to allow the plants to recover from potential detrimental effects of darkness and transportation during shipping (Figure 1).

clear plastic tubes and jars containing plants grown via tissue culture on a black tabletop — Figure 1. Tissue culture sweetpotato plants.

Autoclave commercial potting media and sterilize new 4-inch pots (submerge in 10% bleach solution, rinse with tap water). Place potting media in pots and place in the greenhouse.
After 2-3 days, open the lids of the tubes gradually to equate the relative humidity (RH) with the ambient environment: open lids of the tubes halfway for 2 hours and then fully open for 1 hour. Move plants to the greenhouse.
In the greenhouse, remove the tissue culture plants from the culture tubes and gelling agent by gently pulling them out using forceps (Figure 2). Dip plants in deionized water to remove any traces of solidified agar media (Figure 3). Wear latex gloves disinfected with 70% ethanol to avoid potential contamination of the plants.

a blue gloved hand using forceps holds a tissue culture plant, placing it in a clear plastic tub — Figure 2. Removal of tissue culture plants from culture tubes.

a tissue culture plant that has been cleaned, its stem is green with three tiny leaves and bare white roots — Figure 3. Tissue culture plants after cleaning.

Treat the plants with liquid fungicide(s) (e.g. 2 dithane M-45) by lightly dipping the root base for 2-4 minutes to avoid pathogenic infection.
Insert each plant, holding by the stem, into a pre-formed hole in the potting media, and press potting media lightly around the base for the plant to hold firm and upright.
Place pots inside a humidity box with 25-27 °C and 80-85% RH in the greenhouse (Figure 4). To avoid direct sunlight, cover the humidity box with shaded cloth to maintain 50-70% light. Mist plants and inside of the box before closing the box.

a humidity box sits on a metal greenhouse bench, a clear plastic lid sits over twelve pots with soil and plants, a white wire comes from under the lid and a black wire comes out from the top. — Figure 4. Placing tissue culture plants into a humidity box in the greenhouse.

Gradually open the humidity box by first removing a quarter of the cover after 2-3 days, half on day 4, three-quarter on day 6, and fully uncover on day 8. Monitor the plants daily in the box and mist the plants if needed, but do not water directly.
Place plants on a greenhouse bench and water lightly (Figure 5).
After 3-4 days, place the plants in an open ended hardening-off tunnel for about 1 week (Figure 6).
Transplant the plants to the bed (Figure 7).

a black plastic tray holds 6 black pots with tissue culture plants after a week of growth — Figure 5. Placing tissue culture plants on the green house bench.

a hoop tunnel sits outside near a field. under the tunnel there are many green plants — Figure 6. Plants in open-ended hardening-off tunnel.

tissue culture plants in black plastic pots sit on the black mulch covering the soil in a field row, there are holes in the mulch where the plant will be planted, an orange tag with a variety number sits in the dirt — Figure 7. Tissue culture plants ready to transplant to the bed.

logos for the united stated department of agriculture (left) and the national institute of food and agriculture (right)

The information in this publication is based upon work that is supported by the U.S. Department of Agriculture, National Institute of Food and Agriculture Hatch Project WNP0010 Accession 7005372 and under award number GW24-013 through the Western Sustainable Agriculture Research and Education Program under project number G123-25-WA509. Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and are not meant to endorse any businesses or detract from any not listed.