Effect of Coconut Water (Cocos Nucifera) as an Alternative and Local Source of Plant Growth Hormones and Undefined Nutrient for Ginger Tissue Culture

Effect of Coconut Water (Cocos Nucifera) as an Alternative and Local Source of Plant Growth Hormones and Undefined Nutrient for Ginger Tissue Culture

Ginger is an essential cash crop in Nigeria whose production and propagation is on the decline despite our abundant land and human resources. Ginger production is about 2.5 tons/hectare against the required 3.4 tons/hectare estimated to meet the medicinal and confectionary needs and there has been consistent decline in the production (FAO, 2009).

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Ginger is propagated vegetatively by the rhizome which contributes to the decline in production but tissue culture provides solution to quick propagation to overcome the decline. Tissue culture broadly refers to the in vitro culture of explants under aseptic conditions (sterility) in appropriate nutrient medium under optimum environmental conditions (Ubi, 2013).

Phytohormones are important component of tissue culture which are chemical substances that control growth and developments of plants. However, conventional phytohormones are costly and coconut water is promising to be an alternative and cheap source for economic and availability reasons.

Coconut water (Cocos nucifera) is a clear liquid inside of the fruit of a coconut palm. In early developmental stage, coconut water serves as a suspension for the endosperm of the coconut during their nuclear stage of development. As growth progresses, the endosperm matures into their cellular phase and deposit into the rind of coconut meat which is one of the basic parameters to distinguish between the mature and immature coconut water (Cambel, 2012). In consideration of high content of potassium and other trace metals (e.g. magnesium and phosphorus), low level of fat, mineral content of coconut water, anti-oxidant, L-arginine, ascorbic acid, vitamins, carbohydrates, cytokinins which promote plant cell division and growth and high percentage sterility of coconut water (Rees, 2012), the researcher thought it imperative to research on how effective, and reliable coconut water could be suited for an alternative source of plant growth hormones in tissue culture of ginger in a perfect sterile condition.

AIM:
The aim of this research was:
To determine the effect of concentration, maturity stages, location and varieties of coconut water (Samoan tall and Nias green) on ginger when used as alternative source of phytohormones and undefined nutrient in tissue culture of ginger.

SPECIFIC OBJECTIVES OF THE RESEARCH
The specific objectives of this research include:
1. To compare the growth performance of ginger plants grown in MS medium and MS basal medium supplemented with coconut water via in vitro propagation.

2. To determine the effect of locations of coconut water on tissue culture of ginger when substituted as a phytohormone source.

3. To know the type of coconut water (Samoan tall and Nias green) that is best suited as growth regulator for tissue culture of ginger.

4. To compare and contrast the effect of two maturity stages of coconut water (mature and immature) in tissue culture of ginger when substituted for plants growth hormone.

5. To determine the effect of concentration of coconut water (50 mL, 100 mL and 200 mL) on tissue culture of ginger when used as a source of phytohormone.

 

LITERATURE REVIEW
Coconut Water
Coconut water is the pure liquid found inside of the fruits of coconut palm. It is a suspension for coconut endosperm in the early development stage of coconut. The endosperm progress in to its cellular stage as growth continues and deposits in the rind of coconut meat which is one of the basic parameters to distinguish between the mature and immature coconut water (Cambel, 2012). Coconut water contains high level of potassium, low level of fat, minerals, anti-oxidant, cytokinins (which promote plant cell division and growth), L-arginine, vitamins, carbohydrates, trace metals, (e.g magnesium and phosphorus), and high percentage of sterility (Table 1) (Rees, 2012).

Table 1: Coconut water (Cocos nucifera) fresh nutrition value per 100 g.
Principle Nutrient Value Percentage of RDA
Energy 19 Kcal 1 %
Carbohydrate 3.71 g 3 %
Protein 0.72 g 1.5 %
Total Fat 0.20 g 1 %
Cholesterol 0 mg 0 %
Dietary Fibers 1.1 g 3 %
Vitamins
Folates 3 g 0.75 %
Niacin 0.080 mg 0.5 %
Pantothenic acid .043 mg <1 %
Pyridoxine 0.032 mg 2.5 %
Riboflavin 0.057 mg 4 %
Thiamin 0.030 mg 2.5 %
Vitamin C 2.4 mg 4 %
Vitamin A 0 IU 0 %
Vitamin E 0 mg 0 %
Vitamin K 0 mg 0 %
Electrolytes
Sodium 105 mg 7 %
Potassium 250 % 5 %
Calcium 24 mg 2.4 %
Copper 40 mg 4.5 %
Iron 0.29 mg 3.5 %
Magnesium 25 mg 3.5 %
Manganese 0.142 mg 6 %
Zinc 0. 10 mg 1 %

Human consumption and derivative products
Coconut water (Cocos nucifera), for decades back has been a common drink in tropical countries where they are freshly available, bottled, canned and packaged for human comfortable consumption. Coconut water for drinking is served fresh, packaged in many places or chilled. It is often distributed and sold by street vendors who open them by puncturing with machetes and similar tools in the presence of the consumers. Other processed coconut water for retail can be seen in ordinary cans, or coconut jelly included. Coconut water is also fermented to produce coconut vinegar and also a jelly – like food (Young, 2009).

Commercialization of coconut water
In current years, coconut water is marketed as a natural energy source or sports drink having infinitesimal levels of fat, calories, carbohydrates and significant content of electrolytes. The contents of primary electrolytes per 100 mL serving as unprocessed coconut water potassium, calcium, sodium and magnesium however are insignificant (2-7 % of the daily value) and are not balanced. More so, marketing claims attributing health gains to coconut water are not scientifically based and not allowed by certain regulatory bodies. The Food and Drug Administration (FDA), United States warned producers concerning misleading marketing claims that coconut water is antiviral, regulate blood glucose levels, can lower cholesterol, among other misleading claims, as inappropriate for the product (Cambel, 2000).

Medical uses of Cocos nucifera
Coconut water is rarely used as an intravenous rehydration fluid in place of unavailability of medical saline (Manat, 2012). The history of coconut water appearing similar to human blood plasma evolved during Second World War when Japanese and British patients were administered coconut water intravenously in emergency condition because of unavailability of saline (Manat, 2012). This rehydration technique is implemented for short – term emergency conditions since then in remote areas where plasma is unavailable.

Although using coconut water for saline is not recommended by physicians today, during the Kilmer rouge regime in Cambodian form 1978 to 1979 it was a common practice (Rees, 2012). The documentation centre of Cambodian stated the practice of permitting the administration of coconut water by the untrained nurse during pol- pot regime as a crime against humanity (Rees, 2012). Coconut water can also be used in folk medicine practices in treatment of disease such as diarrhea (Martnez, 2014).

Risk of Excessive Consumption
Anecdotal sources explain coconut water serves for the senicide of aged people, a process called thalaikoothal (Rees, 2012). Customarily, the aged person is given excessive amount of coconut water to drink, which eventually results in fever and subsequent death, the precise cause of which has not been determined. One assumed factor rising from excessive intake of coconut water is excess amount of potassium in the blood (hyperkalemia), causing kidney failure, loss of consciousness, heart arrhythmia and eventually death.

 

Ginger
Ginger (Zingiber officinale) is a perennial flowering plant whose root, rhizome is commonly used as spice and folk medicine. It is herbaceous plant which produces yearly stems of about one meter tall having narrow green leaves and bright yellow flowers. Ginger was first discovered in the tropical rain forest of southern Asia and belongs to the family of zingiberaceae. The ginger plant produced in India exhibits the greatest value of genetic variation and the higher the value of genetic variation the older the plant is assumed to have existed in that region. Ginger was sent from India to Europe in the first century as a result of extensive spice trade and was excessively used by the Romans. Because of their relative taste, the distantly similar dicots in the genus Asarum are generally called wild ginger (Grontled, 2002). Ginger is an ever tropical plant planted for its spicy root (underground stem and rhizomes). The root of a ginger has tan skin, Ivory or pale green flesh, a peppery and fairly pleasant flavor. Ginger is cultivated in August to September. The tops of ginger begin to die prior to flower appearing preparing the roots ready for harvest which takes about 8- 10 months to reach maturity. Ginger is the underground root (rhizome of ginger plant) which has firm texture. Ginger flesh assumes red, yellow or white color based on the variety depending on whether ginger rhizome was harvested when it was young or mature, it is covered with a bright brownish skin that may either be thin or thick.

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Status of ginger production in Nigeria
In 2007, Nigeria had the largest land area of about 195,000 for ginger cultivation globally (FAO, 2009). This stands for approximately 45.4 % of the world total for the year (FAO, 2009). Despite large land area for cultivation, the quantity of ginger produced per unit area is low (Table 2). The estimated quantity of ginger production in Nigeria is about 3.4 tons/hectare in 2009 (FAO, 2009). Under improved and mechanized cultivation conditions, productions as high as 38 tons/hacker could be achieved (Purseglove, 1987). Mechanically, ginger rhizome could be produced in all parts of the country, but in Nigeria it is mostly grown in five states; Nasarawa, Gombe, Kaduna, Niger and Benue. Southern Kaduna is the main producing region with 95 % of the country’s entire production (Okafor, 2002).

Table 2: Area and production of ginger in Nigeria
Year Area (ha) Production (mt)
19998 166,000 88,100
1999 153,000 92,000
2000 158,000 98,000
2001 160,ooo 104,000
2002 162,000 105,000
2003 167,000 110,000
2004 170,000 117,000
2005 181,000 125,000
20006 191,000 134,000
2007 175,000 138,000
Average 170,300 111,110
(Ewuzienm and Nwauzo, 2011). ha = hectare, mt = metric ton.

Chemistry of ginger
The pleasant flavor and aroma of ginger is as a result of the mixture of zingerone, gingerols, shogaols and volatile oils which comprises 1-3 % of the total weight of fresh ginger. In laboratory animals, gingerol raises the motility of gastrointestinal track and instills sedative, analgesic, antipyretic and antibacterial properties. A research at university of michigan showed that gingerols can prevent development of ovarian cancer cell in vitro. The chief pungent principle of ginger is gingerol 1-(4-hydroxy-3-methoxypheny-1) -5- hydroxyl-3- decanone. The non-volatile phenylpropanoid compound, majorly shogaols and gingerols are responsible for the pungent taste of ginger when ginger is cooked or dried. During this process, zingerone is also liberated from gingerols, but less pungent and produces a spicy pleasant aroma (Kim et al., 2008).

Uses of ginger
Ginger gives out a hot, sweet fragrant kitchen aroma. Early ginger rhizomes are freshly and juicy with a cool-mild taste. Ginger is often pickled in vinegar which serves as snacks or barely cooked and added as ingredient in many dishes. Ginger roots are also steeped in hot water to produce ginger tea with the addition of honey, sliced lemon or orange fruits. Commercially, ginger can also be produced into candy or even ginger wine since 1740.
Crystallized ginger or candied ginger is a confectionary from root ginger cooked in sugar until soft. Powdered dried ginger rhizomes are used as an aroma and flavoring for recipes such as cookies, crackers, ginger bread and cakes, ginger beer and ginger ale. Mature ginger roots are fibrous and nearly dry although the aroma and flavor of dried and fresh ginger is partially different. In Korea, China, Japan and South Asia, the fresh juice from ginger rhizomes are usually used as ingredients for flavoring dishes such as various meats, vegetarian cuisine and sea foods.

Composition and safety
Consumption of reasonable amount of ginger has little negative side effect and it is written on the FDA’S “absolutely recognized as safe” list, although it can interact with some medications, even anti-coagulant drug warfarin. Ginger also affects individuals adversely with gall tore. There are also postulations that it can affect clothing, blood pressure and heart rhythms.
Many allergic reactions towards ginger consumption generally leads to rash and though recognized as safe, it can cause heart burn, bloating, nausea, belching precisely if taking in powdered form. People who have ulcers, blocked intestines, inflammatory bowel diseases have a high tendency to react adversely to a large amount of fresh ginger and unmasticated fresh ginger can lead to intestinal blockage in a typical ginger spice serving quantity of 5 g or 1 US table spoon. Ginger powder gives significant quantity of essential nutrient, with the exception of dietary mineral, manganese that is in the daily value quantity of about 79 % (Choudhury, 2000).

Types of ginger
Ginger is commonly mistaken for roots as it yields underground. Botanically speaking, it is a fleshy tuberous rhizome or underground root of a perennial plant comprising of yellow, white and thai ginger as its edible varieties each having different and specific flavor levels of heat. Young edible ginger is always preferred and appreciated for its milder aroma and flavor. Most varieties serve as ornamental plants as a result of their bright lance-shape leaf and their yellow or white flowers and sometimes with violet sparkling color. Ginger can grow up to three feet and very easy to propagate through their rhizomes in ideal tropical climatic environment.

Factually, edible ginger roots left on a covered container can protrude shoots and leaves if delayed. Ginger bears many different names base on the countries they are propagated; Ingwer in German, Zenzero in Italy, and gingermbre in French. It has several names in India where it falls among the most vital agricultural products, it has many names based on maturity, forms and variety. The green variety is known as adruk and dried called sonth.

Baby ginger
In culinary circle, baby ginger is called several names; spring, green, young, stem, new and pink ginger. Baby ginger is milder, softer and more pliant with pink buds. When used as a spice to prepare a dish, whether soup, salad or stir fry, its mild aroma and flavor doesn’t overpower a dish but provides a milder flavor and aroma reminiscent of mature ginger. One of the supposed importance of baby ginger is to eliminate possible parasites and toxins due to its high natural anticeptic properties. Baby ginger is commonly found in Asian markets but very difficult to find in cold region and countries where they are not propagated. Rosy-tipped, small quantity of immature ginger is more mildly aromatic and do not require peeling before cooking. In Asian salad, it is extensively used raw and the Pink-color is appreciated more during its steeping and soaking in vinegar.

Jamaican ginger
The variety of ginger propagated and harvested from Island country of Jamaican is taken as the best available and preferable in the world market. Jamaican ginger is characterized by a woodsy, strong sweet smell. It is harvested all through the year and slicing it yields creamy yellow spice. Like other varieties of ginger, the edible part of Jamaican ginger is planted underground and it is really an improved stem or rhizome. Ginger is still called a root by many circles for culinary reasons. It is quite a home cure for the partial relief of simple cause of motion, vomiting, sickness, dizziness and nausea. In the olden days it served as a temporal food preservative and a natural treatment for digestive problems (Ernest et al., 2000).

Jamaican ginger is regarded by historians from the late 1500s as the first spice from the east, planted in the new world and then imported back to Europe. For domestic uses, Jamaica produces it into flavored ginger beer and other applications include: Jamaican ginger cake and ginger tea produced from the fresh rhizome. Jamaican ginger compared to others grown in other countries is a prized culinary ingredient for its aromatic and pungent qualities.

Organic ginger
Planting organic ginger needs using a piece of organically planted ginger to commence with. Organic ginger does not contain any remnant of potentially harmful compounds and similarly planted free from herbicides, pesticides and synthetic chemical fertilizers. It may not be too critical to use organic ginger for everyday cooking use, since only small quantity is used for most dishes. Using organic ginger for ginger tea, prepared from fresh peeled or unpeeled portion of the mashed root, grantees the body of assimilating only natural healing compounds of the rhizome and not any added artificial chemical potentially remained from conventionally planted ginger. It is preferable to go organic in the choice of ginger that is usually consumed as natural remedy for many illnesses (Marx et al., 2013).

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Planting of ginger
There are two major ways of planting ginger but only one will be discussed: In a pot;
i. Choose and obtain ginger rhizome from a grocery market or store.

ii. Immerse your ginger root in warm water overnight.

iii. arrange a 14 inch (35.6 cm) wide and 12 inch (30.5 cm) deep pot containing potting soil, the loosed type that avoids packing down when watered and enriched with plenty of compost.

iv. Plant the ginger root just below the soil surface spacing the pieces evenly apart.

v. place the pot in an area having light shade at the temperature of 75- 85o F.

vi. Apply water slightly initially and then heavily when shoots appear. Store the plants dry in winter while they are dormant.

vii. Within 10 to 12 months the plant root will be 2- 4 feet and mature.

viii. Uproot new sprout that comes in front of the plant and again replant them somewhere, where they will form new rhizomes

Plant growth hormone
Plant growth hormones are chemical substances that control growth and development of plant. They are otherwise called phytohormones. They are very sensitive molecules produced right inside the plant and are mostly found in virtually low concentrations. Plant regulators control cellular programs in targeted cells where they are manufactured and later transported to other locations of the plant where necessary in order to execute their hormonal functions. Plant hormones determine the flower formation, the height and nature of the leaves, stems, even the appropriate period for leave shading, ripening of the fruits and the entire plant development. Unlike animals, plants lack glands to secrete and produce their vital hormones, rather each cell in every plant is capable of producing its hormones when needed. The general shape of the plant, the nature and size of the seed, flowering period, sex of the flower, leaves senescence, and the fruits are programmed and mastermind by the plant hormones. They orchestrate the down ward growth and upward growth of the plant tissues, piloting the longevity cum death of plant.

Plant hormones also known as plant growth factors are so sacrosanct to plant development and without them plants will end up being masses of undifferentiated cells, with no vital biotechnological formation. Plant hormones are not only seen in multi-cellular plants but also in algae, exhibiting related functions and in similar character, they can also be called secondary metabolites when they are found in micro organisms such as bacteria and fungi where they play no known hormonal and quick physiological functions (Baker et al., 2012).

Characteristics of plant hormones
Phytohormones control transcription levels and gene expression, cellular division, and plant growth. Naturally, they are produced within the plants, serve as chemical substance to regulate and control the growth of plants cultivated and plants grown in vitro. Plant hormones are strictly chemicals, not nutrient and are produced in minute concentrations to influence and promote the development, growth, cells and tissue differentiation. Plants do not have glands to manufacture and keep hormones, but use active passion means to transport their hormones round the plants. The plant hormone biosynthesis is never always localized but is often diffused within the plant tissues. Plants make use of simple substances as hormones that is transported readily and easily within the tissues. They are usually manufactured and utilized on the local basis within plant parts. Plant cells make hormones and control different parts of cells making the hormones.

Plants use four types of movements to transport produced hormones within the plants they are produced. Cytoplasmic streaming around the cells and diffusion of molecules and ions between the cells are used for localized movements. Vascular tissues serve as means to transport plant hormones around the various parts of the plant. These vascular tissues include: Phloem or sieve tubes that transports sugars from leaves to roots and the flowers, the xylem that transports mineral solutes and water from roots to foliage. Not every plant cell responds to hormonal signals, but the cell responds are orchestrated to respond at special and particular points in their life time. The highest effects take place at particular stages during the cells cycle, with drastically reduced effects taking place after or before this specific time.

Plants require hormones at specific periods in plant growth and at particular locations which they need to discard the effect of that hormones when not needed again. Hormone production occurs often at the point of active growth and development around meristems before the complete differentiation of the cells. Different part ways are used by plants to control internal hormone amount and modulate their effects. The quantity of chemicals for hormone biosynthesis is also regulated by plants. If necessary the plant stores hormones in cells, cannibalise formed hormones through the means of conjugating them with peptides, amino acids or carbohydrates or even inactivate them. Plant can effectively destroy hormones by breaking them down chemically but frequently move them from one site to another to dilute their concentrations. The hormonal concentration required by plant for effective response are practically low (10-6 to 10-5 mol/L) (Sathyanarayana, 2007).

Classes of plants hormones
Summary, it is generally accepted that hormones are of five major known classes that are made up of many different chemicals that may differ in structure, varying from one plant to the other. These chemicals are grouped into one of the classes on the bases of their similarity in structure and their physiological effects on plants. Each class exhibits positive and inhibitory functions and most often, works tandemly with each other, with different ratios interplaying to control growth regulation.
Abscisic acid
Abscisic acid (ABA) is an imperative phytohormone that was discovered under different nomenclatures before its real chemical properties were timely known. Abscisic acid was known as abscicin 11 and dormin before it was discovered that both are the same and share the same property and chemical formula. It was given the name abscisic because the highest concentration of the compound was found mostly in newly fallen leaves or freshly abscissed. Abscisic acid is made up of a chemical compound usually manufactured in leaves of plants, coming of chloro plants, mostly during the period of stress. Abscisic acid functions as an inhibitory compound which affects the growth bud, bud dormancy and seed. It controls and mediates effects especially in meristem, producing bud dormancy and alteration of the least terminal leaves to protective buds protector. It functions in seed dormancy and leaf especially in plant species from temperate regions of the world by preventing and inhibiting growth, but immediately the concentration decreases from the buds or seed, growth starts. In many plants, as Abscisic acid concentration reduces, growth begins as the concentration of gibberellins increase. In the absence of abscisic acid seed and buds would commence growth in the period of warm during winter and will die when they froze again.

There is usually a delay in the physiological pathways and routes that offers protection from immature growth because abscisic acid decreases slowly from tissue and its resultant effects consumes time to be offset by other phytohormones. During fruit maturation, abscisic acid gathers within seeds, hindering seed germination in the fruit and seed germination prior to winter. During cold temperature, abscisisc acids effects are reduced in plant tissues liberating the buds and the cells from dormancy, abscisic acid plays many roles in opening and closing stomata in plants undergoing water stress. A signal goes up to the leaves immediately after plant water stressed and the roots undergo water deficiency resulting to abscisic precursors formation there, which then goes to the roots. The roots then liberate abscisic acid that is transferred to the foliage via the vascular system and mediate sodium and potassium up take in the guard cells, which then lose turgidity closing the stomata. It is found in all parts of the plant and the concentration in any tissues mediates its effects and plays a role as a hormone. Its catabolism in the plant effects cellular growth and metabolic reaction and other hormone production. The life of plant commences as seeds with high level of abscisic acid. Prior to the germination of seeds, the concentration of the ABA reduces the more. As the shooting and production of functional leaves commences the level of ABA increases, decreasing cellular development in mature parts of the plants. Abscisic acid catabolism rates and production is affected by predation and stress from water mediating a different cascade of effects that facilitates specific responses from specific cells (Yadav et al., 2009).

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Auxins
Auxins are chemical compounds influencing positively the enlargement of the cells, roots initiation and bud formation. They increase and promote the manufacture of other plant hormones and control stem growth, fruits, roots and transforms stems to flower when combined with cytokinins. Auxins influence elongation of cell by altering the plasticity of cell. They quicken a subclass of meristem called cambium to divide and cause the differentiation of secondary xylem in stem. Auxins functions to inhibit buds growth, lower down the epical dominance and promote adventitious and lateral root growth and development. Auxins contained in seeds regulate certain protein synthesis as they improve after pollination within the flower necessitating flowers to produce fruits to have developing seeds.

In high concentrations, auxins become toxic to plants and the toxicity is more effective to dicots but less to monocots. Synthetic auxin herbicides comprising 2, 4, 5 – T and 2, 4- D are in existence to control weeds because of the toxicity of Auxins in high concentrations. Some classes of auxins more importantly indole-3-butyric acid and 1-nephtylene acetic acid are commonly introduced to stimulate the growth of roots. The auxins and cytokinins (a/c) correlation in plants remain a constant and indole -3- acetic acid is the most popular found auxin.

Cytokinin
Cytokinins are group of chemical compounds that affects division of cells and the formation of shoots and was derived from Zea, discovered in amateur kernels. The first isolation of cytokinins was from yeast cells and was called kinins. Auxin aids tissue delay senescence, responsible for mediating transportation of auxin within the plant and influences leaf growth and intermodal length. In connection to Auxin, Cytokinin has highly synergistic effect and the appropriate ratios of the two phytohormones influence growth period in plant life time. They counter epical dominance caused by auxins and in connection with ethylene improves leaves abscission, fruits and flower parts.

Gibberellins
Gibberellins are large group of chemical compounds naturally produced by fungi and plants. They are vital in germination of seeds, influencing the production of enzymes that orchestrates food production utilized for growth and the development of new cells. Gibberellins achieve this by controlling chromosomal transcription. Water absorption by the seed as a result of layer of cells known as aleurone layer in grain e.g corn, rice, wheat etc leads to gibberellins production. Gibberellins are transported to aleurone layer which works by manufacturing enzymes that catabolize reserved stored foods in the endosperm, which growing and developing seedlings used. Gibberellins manufacture rosette forming bolting of plants improving length of internodes. Gibberellins promote cellular division, flowering and promotes seeds growth after germination. Dormancy and inhibition of shoot growth introduced by abscisic acid is reversed by gibberellin. In summary gibberellins cause stem cell elongation, orchestrates mobilization components of seeds during germination and most importantly in biannials, stimulates bolting and pollen tube growth (Raikhel et al., 2010).

Ethylene
The plant hormone ethylene is a gas produced through the catabolism of methionine that is in every cell. Its solubility in water is very limited and most importantly can’t accumulate in the cell, rather leaves the plant by a way of diffusion. The determinant of the effectiveness of phytohormones is directly proportional to its production verses the rate of diffusion into the atmosphere. In fast dividing and growing cells, especially in darkness, ethylene is rapidly produced. The rate of production of ethylene in newly germinated seedling and new growth is higher than the rate of escape from the plant which results to high concentration of ethylene, preventing leaf expansion.

In plant cells, especially in new shoots phytochrome reactions signal ethylene production to drastically decrease when exposed to the light thereby promoting leaf expansion and development, while underground ethylene influences cell shape and growth when a developing shoot encounters an obstacle, the production of ethylene drastically increases, inhibiting cell elongation, resulting to the swelling of the cell leading to the production of stronger and thicker stem.
The resulting stronger stem can now produce more pressure against the object preventing its parts towards the surface. But if the force exerted by the shoot does not overcome the surface or does not reach the surface, ethylene stimulation and production becomes prolonged, the effects affects natural geotropism of the stem, which should grow up right, controlling it to grow within an object. Experiments carried out shows that ethylene influences stem height and diameter. High concentration of ethylene production occur when the tree stems are subjected to whirl wind resulting to lateral stress leading to thicker, stronger sturdy tree branches and truck. Ethylene influence fruit ripening: usually, during maturation of seed ethylene production enhances and accumulate in the fruit leading to climacteric activities before seed dispersal. Ethylene production also regulates nuclear protein ethylene insensitive 2 and regulates other phytohormones including stress hormones and ABA (Trigiano et al., 2000).

Plant tissue culture
Plant tissue culture broadly refers to the in vitro culture of explants under aseptic conditions (sterility) in appropriate nutrient medium under optimum environmental condition (Ubi, 2013). For the sake of this study, tissue culture of ginger refers to the in vitro culture of ginger explant under sterile condition in appropriate nutrient medium under optimum environmental condition. It is usually carried out via use of the liquid, semi-solid or solid growth media, such as broth or agar (White, 1993). Tissue culture techniques and procedures will be strictly used to culture ginger using coconut water as the major source of plant growth hormone and undefined nutrient in this research.

Tissue culture procedures
Choosing and preparing a suitable nutrient media.
Choosing explant (ginger) from a healthy, vigorous mother plant.
Surface sterilization of the selected explant by disinfectants.

Transfer of the explants on the suitable sterile nutrient medium in culture vessels under laminar flow hood sterile conditions.
Growing the cultures in a controlled growth chamber at optimum conditions of light (16 hours of photoperiod), diurnal illumination, temperature (25+2o C) and 70-75 % relative humidity.
Regeneration of whole plants from cultured plant tissue.

Hardening
Transfer of the acclimatized plants to the field.

Modern usage of tissues culture
In modern application and usage, tissues culture refers to growth of cells form a tissue of a multicelluar organism in vitro. The cell can be cells isolated from a primary cells donor organism, or an immortalized cell line. These cells are bathed in culture nutrient medium that contains important nutrients and source of energy necessary for the cells survival. Tissues culture is often used interchangeably with cell culture. The practical meaning of tissues refers to the culturing of tissue pieces that is explants culture.

Tissue culture is an essential tool for the learning of biology of cells from multicellular organism. It makes available an in vitro model of the tissue in a perfect sterile environment that can be easily manipulated and analyzed. Plant tissue culture itself is particularly concerned with the growing of total plants from a small piece of plant tissues, maintained and cultured in a medium.

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