Acclimatization of MD2 Pineapples in Nursery

Acclimatization of MD2 Pineapples in Nursery

The in vitro plantlet from tissue culture lab is not suitable to be planted immediately in the field. A physiological adaptation process called acclimatization is carried out in the nursery with a serial of hardening treatments to ensure the plantlets adapted through the stressful external environment before it can be planted in the field.

The ex-lab pineapple plantlets were transferred into the polybag containing planting substrates and supplemented with nutrients in order able to acclimatize through this transition period in the nursery. Under the monitored temperature, humidity and light, the plantlets developed into hardened seedlings after few weeks in the green house. During the acclimatization process, the leaf tissue of plantlet will grow thicker it will strengthen the whole leaf structure. An extracellular hydrophobic layer is also developed to cover aerial epidermis of plantlets providing protection against desiccation and external environmental stresses.

Gain Green Development’s laboratory offers high-quality tissue cultures to fulfil the customer’s demand. By Company’s team experts in keeping up protocols and creating new ones for untested plant species and varieties.

Contact us for further discussion!

Thai black galingale (Kaempferia parviflora Wall. ex Baker)

Kaempferia parviflora Wall. ex Baker, also known as Thai black galingale, Thai ginseng or Cekur Hitam in Malay and Krachaidam in Thailand, is an important medicinal plant from the Zingiberaceae family. K. parviflora is a perennial and herbaceous stemless plant with dark purple rhizomes with a taste that has spiciness to it, but much more subdued than ginger. It is originally found in the North and Northeast of Thailand. The leaves of K. parviflora is approximately 6 to 8 cm long, oblong to lanceolate in shape with red margins, and produces purple and white flowers (Labrooy et al., 2013; Labrooy et al., 2020). In the Zingiberaceae family, Zingiber officinale and Curcuma longa are commonly used in culinary to enhance the taste and aroma of the dishes.

Nowadays, natural product derived medicine is preferable to modern medicine because of its low side effects. K. parviflora is widely used as an alternative medicine in treating various types of diseases including fungal infections, gastrointestinal disorders, and decreased vitality and allergies (Tewtrakul et al., 2008; Trisomboon, 2009). In several studies conducted on K. parviflora, the extract of this plant has potential as an anti-inflammatory, anti-allergies, anti-cancer, anti-plasmodial, anti-fungal, anti-HIV-1 protease activity, and antispasmodic effects (Sookkongwaree et al., 2006; Wattanapitayakul et al., 2008; Sae-wong et al., 2009; Saokaew et al., 2016). It is known as health-promoting herbs and traditionally used as a folk medicine for managing a variety of diseases, including ulcers, gout, colic disorder, abscesses, and osteoarthritis.

This phenomenon has led to the demand for medicinal plants including K. parviflora, to increase drastically. This plant undergoes a vegetative stage for three months and a reproductive stage for two months. This plant produces flowers, but the flowers are inconspicuous and do not produce seeds (Labrooy et al., 2020). The long dormancy period and the inability to set seed affect raw material production for K. parviflora. However, the demand for K. parviflora rhizomes in Malaysia can hardly be fulfilled due to the scarcity of planting materials (Labrooy et al., 2013). Propagation of K. parviflora via conventional propagation is time consuming due to the long dormancy period after senescence, which is approximately 5 to 7 months during November to May according to the Malaysian climate (Techaprasan et al., 2010). To overcome the limited supply of K. parviflora raw materials, plant tissue culture technique plays a pivotal role in cloning the black ginger at mass scale.


Labrooy, C., Thohirah, L. A., Johnson, S., Nur Ashikin, P. A. and Maheran, A. A. (2013). Morphological description for kunyit hitam (Kaempferia parviflora) and breaking bud dormancy with BAP and Ethephon treatment. Transactions of the Malaysian Society of Plant Physiology, 22, 139-141.

Labrooy, C., Abdullah, T. L. and Stanslas, J. (2020). Influence of N6-Benzyladenine and sucrose on in vitro direct regeneration and microrhizome induction of Kaempferia parviflora Wall. Ex Baker, an important ethnomedicinal herb of Asia. Tropical Life Sciences Research, 31(1), 123-139.

Sae-wong, C., Tansakul, P. and Tewtrakul, S. (2009). Anti-inflammatory mechanism of Kaempferia parviflora in murine macrophage cells (RAW 264.7) and in experimental animals. Journal of Ethnopharmacology, 124(3), 576-580.

Saokaew, S., Wilairat, P., Raktanyakan, P., Dilokthornsakul, P., Dhippayom, T., Kongkaew, C. and Chaiyakunapruk, N. (2016). Clinical effects of krachaidum (Kaempferia parviflora): A systematic review. Journal of Evidence-Based Complementary and Alternative Medicine, 22(3), 413-428.

Sookkongwaree, K., Geitmann, M., Roengsumran, S., Petsom, A. and Danielson, U. H. (2006). Inhibition of viral proteases by Zingiberaceae extracts and flavones isolated from Kaempferia parviflora. Die Pharmazie-An International Journal of Pharmaceutical Sciences, 61(8), 717-721.

Techaprasan, J., Klinbunga, S., Ngamriabsakul, C. and Jenjittikul, T. (2010). Genetic variation of Kaempferia (Zingiberaceae) in Thailand based on chloroplast DNA (psbA-trnH and petA-psbJ) sequences. Genetics and Molecular Research, 9(4), 1957-1973.

Tewtrakul, S., Subhadhirasakul, S. and Kummee, S. (2008). Anti-allergic activity of compounds from Kaempferia parviflora. Journal of Ethnopharmacology, 116(1), 191-193.

Trisomboon, H. (2009). Kaempferia parviflora, a Thai herbal plant, neither promote reproductive function nor increase libido via male hormone. Thai Journal of Physiological Sciences, 21, 83-86.

Wattanapitayakul, S. K., Chularojmontri, L., Herunsalee, A., Charuchongkolwongse, S. and Chansuvanich, N. (2008). Vasorelaxation and antispasmodic effects of Kaempferia parviflora ethanolic extract in isolated rat organ studies. Fitoterapia, 79(3), 214-216.


Ginger (Zingiber officinale Roscoe)

Medicinal plants have long been used in the treatment of several diseases throughout the world. Ginger (Zingiber officinale Roscoe), a herbaceous perennial plant from the Zingiberaceae family, is one of those plants. There are 24 genera and 300 species under this family; the genus Zingiber has about 20 species as well (Newman, 2001). It is found in tropical and subtropical regions that bear flowers. It used as a spice, food, flavouring agent, and medicine.

In 2019, the global production of ginger was 4.08 million tons, which demonstrates its significant economic value in world trade (Li et al. 2021). It cannot be sexually propagated due to poor flowering and ginger is an unfertile species that failed to set seed (Kambaska & Santilata, 2009). Therefore, ginger plant possesses perennial tuberous or rhizomatous that are used for its vegetative propagation (Nair, 2019). The plant generates an upright, annual stalk (pseudo-stem), 60 to 90 cm tall, with dark green leaves. Its stalks are covered with flat sheaths that may be taken off stalk; 8 – 12 distiches leaves are present on the stem. The leaves are with long blades, or flat and stalk less blades; are alternative (alternate), lance late, linear lance late, specula, 10 to 21 cm tall and 2 to 2.5 cm wide.

The presence of a high polyphenols and flavonoids content in its leaves, stem, and rhizome has been defined as the critical factor for its pharmacological effects (Ghasemzadeh et al., 2010). These polyphenols and flavonoids compounds are natural sources of antioxidants (Haida et al., 2019). As rhizome is an economically exploited part of the plant, using a high proportion of ginger rhizomes as starting material for cultivating the plant in the next growing season negatively affects its supply in the market. It has been used as spice and medicine for treating cancer (Zhang et al., 2021); cardiovascular disease (Ghafoor et al., 2020); diabetes (Said et al., 2020); and several other illnesses such as cold, nausea, asthma, and cough (Choi et al., 2018). Owing to the global pandemic of COVID-19, ginger consumption gained more interest. It helped alleviate the severe symptoms of COVID-19 positive patients and reduced the recovery time in those patients (Rangnekar et al., 2020).

The success of in vitro technique largely depends on the aseptic culture establishment, shoot regeneration capacity, rooting, and acclimatization. Rhizome buds, which are often used as the source of explants in Zingiberaceae, have been proven to be more responsive. However, the initial establishment of contamination-free culture is difficult owing to the exposure of rhizomes to various soil pathogens (Meenu & Kaushal, 2017; Thakur et al., 2018). These pathogens need to be eliminated by surface sterilization of the explants. Vegetative propagation of ginger has a high risk of spreading infections. Slow propagation rate and the risk of disease transmission by sectioning of the rhizomes have deprived propagation by conventional means. Therefore, plant tissue culture is considered the best alternatives method that may supply a large number of planting materials (Hamirah et al., 2010).


Choi, J.G.; Kim, S.Y.; Jeong, M.; Oh, M.S. Pharmacotherapeutic potential of ginger and its compounds in age-related neurological disorders. Pharmacol. Ther. 2018, 182, 56–69.

Ghafoor, B.; Ali, M.N.; Riaz, Z. Synthesis and appraisal of natural drug-polymer-based matrices relevant to the application of drug-eluting coronary stent coatings. Cardiol. Res. Pract. 2020, 2020, 1–11.

Ghasemzadeh, A.; Jaafar, H.Z.E.; Rahmat, A. Antioxidant activities, total phenolics and flavonoids content in two varieties of Malaysia young ginger (Zingiber officinale Roscoe). Molecules 2010, 15, 4324–4333.

Haida, Z.; Syahida, A.; Ariff, S.M.; Maziah, M.; Hakiman, M. Factors affecting cell biomass and flavonoid production of Ficus deltoidea var. kunstleri in cell suspension culture system. Sci. Rep. 2019, 9, 9533.

Hamirah, M. N., Sani, H. B., Boyce, P. C., and Sim, S. L. 2010. Micropropagation of red ginger (Zingiber montanum Koenig), a medicinal plant. J. Mol. Biol. Biotechnol., 18(1): 127-130.

Kambaska, K. B., and Santilata, S. 2009. Effect of plant growth regulator on micropropagtion of Ginger (Zingiber officinale Rosc.) cv- Suprava and Suruchi. J Agric Technol., 5(2): 271-280.

Meenu, G.; Kaushal, M. Diseases infecting ginger (Zingiber officinale Roscoe): A aeview. Agric. Rev. 2017, 38, 15–28.

Thakur, M.; Sharma, V.; Kumari, G. In vitro production of disease free planting material of ginger (Zingiber officinale Rosc.)—A single step procedure. Res. J. Biotechnol. 2018, 13, 25–29.

Li HL, Wu L, Dong Z et al (2021) Haplotype-resolved genome of diploid ginger (Zingiber officinale) and its unique gingerol biosynthetic pathway. Hortic Res.

Nair, K.P. Turmeric (Curcuma longa L.) and Ginger (Zingiber officinale Rosc.)-World’s Invaluable Medicinal Spices: The Agronomy and Economy of Turmeric and Ginger; Springer Nature: Basel, Switzerland, 2019; ISBN 9783030291884.

Newman, M. 2001. Nomenclatural notes on Zingiberaceae. J. Bot., 58(1): 173-174.

Rangnekar, H.; Patankar, S.; Suryawanshi, K.; Soni, P. Safety and efficacy of herbal extracts to restore respiratory health and improve innate immunity in COVID-19 positive patients with mild to moderate severity: A structured summary of a study protocol for a randomised controlled trial. Trials 2020, 21, 943.

Said, H.; Abdelaziz, H.; Abd Elhaliem, N.; Elsherif, S. A comparative study between ginger and Echinacea possible effect on the albino rat spleen of experimentally induced diabetes. Egypt. J. Histol. 2020, 43, 763–776.

Zhang, M.M.; Wang, D.; Lu, F.; Zhao, R.; Ye, X.; He, L.; Ai, L.; Wu, C.J. Identification of the active substances and mechanisms of ginger for the treatment of colon cancer based on network pharmacology and molecular docking. BioData Min. 2021, 14, 1–16.


Ananas comosus MD2 pineapple

Pineapple (Ananas comosus) is one of the most economically important tropical fruits within the Bromeliaceae family (Duval et al., 2001). It is the most highly traded tropical fruit. Pineapple is a perennial herbaceous and monocotyledonous flowering plant whereby it sends up a flower stalk from that central point. Generally, it is usually asymptomatic and has a range of height and width of 0.91 to 1.98 m (Bartholomew et al., 2003). In Malaysia, we have many varieties, namely Moris (AC1), Sarawak (AC2), Gandol (AC3), Maspine (AC4), Josapine (AC5), Yankee (AC6), Moris Gajah (AC7), N36 (AC8), MD2 (AC9), and many more. Among all, MD2 is the most favourite variety due to its super sweet content and yellowish flesh that commonly involved in food and beverage industry.

Top pineapple producing country is Costa Rica, followed by Philippines and Brazil. Malaysia produces about 330k tons annually. For the Southeast Asian region, Indonesia is among the third largest pineapple producers after the Philippines and Thailand with a contribution of around 23%. Pineapple is mostly consumed worldwide as fresh fruit due to its rich flavour, taste, and size. Consumed this way, it can be an excellent source of vitamin A, C, vitamin B1, vitamin B6, manganese, magnesium, iron, copper, phosphorus, and dietary fibre (“Fresh Pineapples,” 2008). It has no starch and consumers only need to peel, core, and slice the pineapple for consumption. Canned pineapple has a wide market as processed fruit. There are a great variety of the canned presentations such as whole, slices, or rings, fingers or spears, cubes, chunks, wedges, and tidbits.

Plain cans are used to pack the fruit because they do not contain tin plate which dissolves due to the fruit acid (Lobo & Paull, 2017). Sometimes the fruit is packed in syrup to hold its colour, shape, and flavour or it can be canned with water or fruit juice to reduce the sugar content. Other downstream products can be made from the pineapples are dried fruits, frozen pineapple, pineapple juice, pineapple jam, and meat tenderizer (bromelain). Pineapple leaf fibre is a hair-like material in elongated pieces similar to thread. The leaf fibre is a comprised mainly of cellulose (Bongarde & Shinde, 2014).

The seeds of these plants are very slow to germinate and therefore are not used for commercial purposes. This bromeliad is routinely propagated vegetatively by means of various vegetative parts, organs, or tissues such as:

  1. basal suckers originating from buds below ground level,

  2. hapas, which are shoots developing at the base of the peduncle,

  3. leafy branches arising from buds in leaf axils,

  4. slips, which grow out of the peduncle below or at the base of fruit,

  5. crowns, arising from the upper part of the fruit, and

  6. butts or stumps from the mature plant (Rangon, 1984).

In case of industrial scale production suckers and heaps are also used (Firoozabady and Gutterson, 2003). The tissue culture method of pineapple comes into the play to increase the selectivity of the desired traits coupled with a high multiplication rate. Following the standard tissue culture method (Mathews and Rangan, 1979; Zepeda and Segawa, 1981; Fitchet, 1990; Fitchet-Purnell, 1993; Kiss et al., 1995; Gangopadhyay et al., 2005) much higher rate of multiplication (40 to 85 fold in a 13 month period) was obtained.

Pineapple crops contaminated with viruses affect production and may lead to mealybug wilt of pineapple (MWP) disease which can be found in all pineapple crops in high density growing areas throughout the world (Sether et al. 2001). Mealybug wilt of pineapple symptoms are morphologically evident on plant leaves displaying leaf downward curling, reddening, wilting, or gradual dying of the tips (Sether et al., 2001; Sether et al., 2005).

Plant tissue culture is about one of the major option to develop an efficient and economical micropropagation protocol for the large scale propagation of pineapples. The micropropagation protocol has now become fairly standardized for some important cultivars of pineapple and with minor changes can be applied to newly derived ones. Propagation in temporary immersion bioreactors (Espinosa et al., 2002) which claims to be an improvement over the existing micropropagation protocol, reports a better performance of such plantlets over those propagated by conventional methods of micropropagation.


Bartholomew, D., Paull, R., & Rohrbach, K. (2003). The pineapple (pp. 1-291). CABI Publishing.

Bongarde, U. S., & Shinde, V. D. (2014). Review on natural fiber reinforcement polymer composties, 3.

Center for Agricultural Data and Information Systems Secretariat General of the Ministry of Agriculture 2016 Agricultural Commodities, Horticulture Sub Sector: Pineapple Outlook accessed on December 18th 2019

Duval MF, Noyer JL, Perrier X, Coppens d’Eeckenbrugge G, Hamon P (2001). Molecular diversity in pineapple assessed by RFLP markers. Theor. Appl. Genet. 102: 83-90.

Espinosa, P., Lorenzo, J.C., Iglesias, A., Yabor, L., Menendez, E., Borroto, J., Hernandez, L. & Arencibia, A.D. (2002) Production of pineapple transgenic plants assisted by temporary immersion bioreactors. Plant Cell Rep. 21, 136–140.

Firoozabady, E. and Gutterson, N. 2003. Cost effective in vitro propagation method for pineapple. Plant Cell Reproduction. 21: 844-850.

Fitchet, M. 1990. Clonal propagation of Queen and Smooth Cayenne pineapple. Acta Horticulturae. 275:261-266.

Fitchet-Purnell, M. 1993. Maximum utilization of pineapple crowns for micropropagation. Acta Horticulturae. 334: 325-330.

Fresh Pineapples. (2008). Retrieved April 1, 2017, from

Gangopadhyay, G., Bandyopadhyay, T., Poddar, R., Basu Gangopadhyay, S. and Mukherjee, K.K. 2005. Encapsulation of pineapple microshoots in alginate beads for temporary storage. Current Science. 88: 972-977.

Kiss, E., Kiss, J., Gyulai, G. and Heszky, L.E. 1995. A novel method for rapid micropropagation of pineapple. Horticulture Science. 30: 127-129.

Lobo, M. G., & Paull, R. E. (2017). Handbook of Pineapple Technology: Production, Postharvest Science, Processing and Nutrition. Wiley.

Mathews, H.V. and Rangan, T.S. 1979. Multiple plantlets in lateral bud and leaf explant in vitro cultures of pineapple. Scientia Horticlturae. 11: 319-328.

Rangan, T.S. 1984. Pineapple, p. 373–382. In: P.V. Ammirato, D.A. Evans, W.R. Sharp, and Y. Yamada (eds.). Handbook of plant cell culture 3, Crop species. Macmillan, New York.

Sether, D., A. Karasev, C. Okumura, C. Arakawa, F. Zee, M. Kislan, J. Busto, and J. Hu. 2001. Differentiation, Distribution, and Elimination of Two Different Pineapple mealybug wiltassociated viruses Found in Pineapple. Plant Disease 85:856-864.

Sether, D., M. Melzer, J. Busto, F. Zee, and J. Hu. 2005. Diversity and Mealybug Transmissibility of Ampeloviruses in Pineapple. Plant Disease 89:450-456.

Zepeda, C. and Sagawa, Y. 1981. In vitro propagation of Pineapple. Horticulture Science. 16:495.