Completing the production process in vitro on hana strawberries and investigating the control factor for strawberry flowering

Effects of AC on rooting ability of strawberry shoots after 3 weeks 38 Table 4.7.. Effects of α-NAA on rooting ability of strawberry shoots after 3 weeks .... 39 Table 4.8.Effects of IBA

VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE

FACULTY OF BIOTECHNOLOGY

GRADUATE THESIS

COMPLETING THE PRODUCTION PROCESS IN VITRO ON

HANA STRAWBERRIES AND INVESTIGATING THE CONTROL FACTOR FOR STRAWBERRY FLOWERING

Hanoi - 2021

VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE

FACULTY OF BIOTECHNOLOGY

GRADUATE THESIS

COMPLETING THE PRODUCTION PROCESS IN VITRO ON

HANA STRAWBERRIES AND INVESTIGATING THE CONTROL FACTOR FOR STRAWBERRY FLOWERING

Practicing student Mao Thi Thuy Trang

Hanoi - 2021

ACKNOWLEDGEMENT

This study would not have been possible without the support of many people I would like to express my gratitude to my professors of Department of Plant Biotechnology at Faculty of Biotechnology in Vietnam National University of Agriculture for their support and conducting the research Particular thanks to Nguyen Thanh Hai, Ph.D for his valuable comments and helpful advice, scientific guidance and valuable feedback throughout the time at laboratory of Department of Plant Biotechnology, especially with me work

Finally, I wish to express my special thanks to my family for their great encouragement throughout my project and thesis, as well as my friends for their continual supports

Table of Contents

COMMITMENT i

ACKNOWLEDGEMENT ii

ABBREVIATIONS v

LIST OF TABLES vi

LIST OF FIGURE vii

I INTRODUCTION 1

1.1 Problem 1

1.2 Purpose and requirement 2

1.2.1 Purpose 2

1.2.2 Requirement 2

1.3 Research location 2

II LITERATURE REVIEW 3

2.1 General introduction 3

2.2 Origin 3

2.3 Distribution 4

2.4 Category 4

2.5 Botanical characteristics 5

2.6 Strawberry diseases 6

2.6.1 Causes of the disease: 6

2.6.2 The most common strawberry diseases 7

2.7 Nutritional value and economic value 18

2.8 Factors regulating development 20

2.9 Research in the world and in Vietnam 22

III Materials and experiments 25

3.1 Material 25

3.2 Experiments 25

3.2.1 Production process of in vitro strawberry 25

3.2.2 Survey of plant physiological properties controlling strawberry flowering

27

3.2.3 Rooting 27

IV EXPERIMENT COMPLETED 28

V CONCLUSIONS AND SUGGESTIONS 42

VI REFERENCES 43

ABBREVIATIONS

LIST OF TABLES

Table 4.1 Influence of disinfectant to the stage to create the initial materials 30

Table 4.2 Effect of BA on the multiplication of strawberry after 5 weeks 31

Table 4.3 Effect of BA and Glutamine on the multiplication of strawberry after

5 weeks 33 Table 4.4 Effect of BA and coconut water on the multiplication of strawberry after 5 weeks 35 Table 4.5 Effect of Reduced lighting hours on the flowering of strawberry after

4 weeks 37 Table 4.6 Effects of AC on rooting ability of strawberry shoots after 3 weeks 38 Table 4.7 Effects of α-NAA on rooting ability of strawberry shoots after 3 weeks 39 Table 4.8.Effects of IBA on rooting ability of strawberry shoots after 3 weeks 41

LIST OF FIGURE

Figure 2.1 Hemorrhoids on strawberries 7

Figure 2.2 Helix aspersa Slug on strawberries 9

Figure 2.3 White powder disease on leaf 10

Figure 2.4 White chalk disease on strawberry plants 10

Figure 2.5 Rubber disease 11

Figure 2.6 Black spot disease 12

Figure 2.7 Gray mold on strawberry plants 13

Figure 2.8 Gray mold on strawberry diseases 14

Figure 2.9 Blackberry root rot 15

Figure 2.10 Red Spider 16

Figure 2.11 Tetranycus Urticae red spider 18

Figure 4.1 The shoots of strawberry create the initial materials 28

Figure 4.2 Strawberry shoots in vitro after sterilization on background medium after 4 weeks 30

Figure 4.3 Strawberry shoots in vitro on basic medium supplemented with BA after 5 weeks 32

Figure 4.4 Strawberry shoots in vitro on basic medium supplemented with BA and Glutamine after 5 weeks 34

Figure 4.5 Strawberry shoots in vitro on basic medium supplemented with BA and Coconut water after 5 weeks 36

Figure 4.6 The shoots young of strawberry in lighting hours is 10h 37

Figure 4.7 Roots of strawberry shoots on basic medium supplemented with AC after 3 weeks 39

Figure 4.8 Roots of strawberry shoots on basic medium supplemented with α-NAA after 3 weeks 40

Figure 4.9 Roots of strawberry shoots on basic medium supplemented with IBA after 3 weeks 41

I INTRODUCTION

1.1 Problem

The strawberry is a perennial which arises from a crown of meristematic tissue or compressed stem tissue Leaves, stems, runners, axillary crowns, inflorescences, and roots all arise from the crown The plant has trifoliate leaves which spiral around the crown, with buds in the leaf axils giving rise to the runners Runners have two nodes with a plant produced at the distal node Strawberry blossoms contain many pistils, each with its own style and stigma attached to the receptacle When fertilization occurs the receptacle develops into

a fleshy fruit (Darnell, R., 2003)

The fruit is called an achene which contains the seeds The edible part is an accessory type fruit The seeds are arranged on the outside of the receptacle tissue The growth of the receptacle is dependent on successful fertilization of the ovules with its size and shape dependent on the number of achenes formed Strawberry plants are day length dependent with cultivars being long day, short day or day neutral (Darnell, R., 2003)

Following further hybridizations, especially since 1850, strawberries has developed into the large, fragrant, tasty red fruit that is now cultivated worldwide The high degree of genetic heterozygosity present in Fragaria spp enabled the development of strawberry cultivars adapted to widely varying environment conditions and resistant to several diseases and pests Not only the genetic variability, but also a high adaptability and plasticity of the strawberry plant itself give this crop such a remarkable range of adaptation (Darrow, G.M., 1966)

That heterozygosity was explained by (Gaafar and Saker 2006) as there are more than 20 Fragaria species worldwide, there are seven basic types of chromosomes that they all have in common However, they exhibit different polyploidy Some species are diploid, having two sets of the seven chromosomes (14 chromosomes total) Others are tetraploid (4x = 28),

hexaploid (6x = 42), octoploid (8x = 56) or decaploid (10x = 70) (Emarah.,

- Redefine the appropriate fast multiplication environment

- Redefine the medium to create suitable complete roots

- Find the suitable lighting time to increase yield and prolong the harvest of strawberry

1.3 Research location

- The study was performed at Department of Plant Biotechnology, Faculty

of Biotechnology and Institute of Agro- Biology in Vietnam Nation University

of Agriculture

II LITERATURE REVIEW

2.1 General introduction

The strawberry - Fragaria is scientifically known, is a highly economical

crop It’s a fruit with very high nutritional value, contain a lot of vitamin C, malic acid, rich in fiber, antioxidants This fruit has many benefits for human health and provides a relatively high source of income for strawberry growers

2.2 Origin

Strawberries ( scientific name : Fragaria ), also known as strawberry is a

genus of flowering plants and species of flowering plants belonging

to Rosaceae ( Rosaceae ) for fruits are more popular Strawberries originated in

the Americas and were bred by European gardeners in the 18th century to create

a widely cultivated strawberry variety today This species was first scientifically

described by (Weston) Duchesne in 1788

In 2012, a Japanese guy named Nahana Shojiro decided to live in Vietnam

and chose Moc Chau as a stopover land He brought along the famous Tochiotome strawberry from the famous Tochigi region After bringing Tochiotme strawberry varieties planted experimentally in Moc Chau, after a period of living and working hard with passion and rightly, he chose Moc Chau

to stop Japanese strawberry variety has established a certain brand in this plateau

Tochiotome strawberry variety has gradually been adopted by consumers

by planting on Moc Chau land on sunny days, cold nights, making Japanese strawberries much sweeter than strawberries Da Lat or Korea The deep sweetness of the Tochiotome Japanese strawberry variety is very suitable for the taste of Vietnamese consumers Because this strawberry variety has a long Japanese name that makes it difficult for Moc Chau and Son La people to call,

then they took the name of Japanese farmer Nahana Shojiro to name this strawberry variety, short for easy Remember "Hana Strawberry”

2.3 Distribution

Strawberries adapt to a variety of ecology: temperate, Mediterranean, subtropical and sub-temperate Around the world, strawberries are grown in many countries such as the US, France, Japan, the Netherlands

Currently, Tochiotome (Hana) of Japan has been propagated and grown

popularly in Moc Chau, Son La as well as many provinces from North to South

2.4 Category

There are over 20 different varieties of strawberries around the world The key to classifying strawberries is based on their number of chromosomes There

are 7 basic types of chromosomes that all have in common However, they

exhibit different polyploidy Some species are diploid, having 2 sets containing

7 chromosomes (2n = 14) Other species are tetraploids (4 sets, 4n = 28), hexadecimal (6 sets, 6n = 42), octal (8 sets, 8n = 56) or multiple (10 sets, 10n = 70)

As a rule of thumb (with some exceptions), strawberries with more chromosomes tend to produce larger, healthier plants with larger berries (according to Darrow)

Fragaria orientalis

Hexaploid

Fragaria moschata (muskberry)

Bat multiples and hybrid

Fragaria x ananassa (Garden strawberry)

Fragaria chiloensis (Chilean strawberry)

Fragaria iturupensis (Iturup Strawberry)

Fragaria virginiana (Virginia Strawberry)

Cross and hybrid

Leaves: Leaves have the shape, structure, thickness and amount of undercoat, depending on the variety Most strawberry varieties have double leaves with 3 leaflets, some varieties have doublet leaves with 4 or 5 leaflets The edges are serrated The petiole is long, the petiole is usually white when the leaves are young, and turns red when the leaves are old

Flower: Divided into many branches, each branch has a flower The flower has 5 thin, white, slightly rounded petals Flowers are hermaphrodite, have 25-

30 stamens and 50-500 stamens Strawberries are fertile but through pollination to increase the frequency of desired genes and to produce some species

self-Fruit: is a kind of fake fruit due to the bulging flower base, it is indeed located on the outside of the fake fruit The fruit is oval in shape; the young fruit

is green When the fruit is ripe, the fruit is pink or red depending on the

variety Strawberries are fragrant, sweet and sour (Lucy, 2017)

Roots: A root system, roots develop about 30 cm from the ground

2.6 Strawberry diseases

2.6.1 Causes of the disease:

Weather: If it is cold, the stamens will die leading to brown flowers, if the weather is too cold, the flower will die, some of the flowers that have been pollinated will survive and make the flower deform In the cold season, when the temperature is too low, the cells are frozen, the base is browned, the plant is poorly grown, and is susceptible to pests and diseases

Light intensity: The light intensity is high, the cells will die, there are circular marks on the leaves

Hail: Hail causes leaves, flowers and fruit to be crushed, creating

opportunities for pathogens to penetrate, greatly damaged plants, causing brown

marks on leaves due to scratches

- Nutritional disorders:

+ Nitrogen: Initially, the plant needs a great amount of nitrogen If there is

a lack of protein leaves, the fruit will be small, the tree will give a little look, the old leaves turn orange or red, the young young and pale green Excess nitrogen

Analyzing soil samples and based on growth and symptoms of plants to adjust nitrogen accordingly

+ Potassium: Potassium- deficient plants are wilted, old leaves are dry, and fruits are easy to rot

Using potassium bicarbonate sprayed on can prevent disease and provide more potassium to the plant; Regularly supply potassium fertilizers to plants such as KNO3, K2SO4

+ Boron: Lack of Bo is one of the reasons that lead to smaller and deformed mulberry fruit, as some areas on the fruit shrink and fail to develop The taste of the fruit is almost normal, but of no economic value

Bo is very important in the pollination process When one or more female flowers are not pollinated, the fruit tissue there will not grow and deform the fruit Boron has an important function for the root system, so a deficiency in boron can prevent the mulberry plant from absorbing nutrients well

+ Calcium: When the leaves are young and not yet spread, the ends of the leaves are necrotic and dry, so when the leaves grow, the tips are twisted but other parts of the leaves are still developing normally and healthy Strawberry plants that grow too fast often interfere with the absorption of calcium from the soil, although the calcium in the soil is abundant The climate is dry, cold, and cloudy, which greatly impedes the plant's calcium absorption

+ Herbicides: Using improper herbicide, the correct dose of plant will die, selective herbicide, pre-germination or post-germination herbicide

2.6.2 The most common strawberry diseases

2.6.2.1 Hemorrhoids: Thrip tabaci

Figure 2.1 Hemorrhoids on strawberries

Morphological characteristics: mature small, yellow-gray color, eggs are laid in tissue in young parts of plants, female thrips can lay eggs 40-50 eggs Young thrips are pale yellow in color, and live harmlessly with adult thrips The thrips is an intermediate metamorphic insect, the young thrips that go into a pseudo-pupal stage that can be in dry leaves or bark, but mostly still in the

soil The life cycle of thrips is 17-20 days, a year there can be about 20 generations of thrips complete development cycle

Harmful characteristics arise law: Hemorrhoids damage mainly on flowers, making the fruit small and deformed In addition, they also damage leaves, young buds and stems, sting and absorbing sap, making plants exhausted, reducing harvest yield Harmful flowers turn brown However, young fruit continues to grow but has a golden color

Fruits with these symptoms are usually small and hard, and the seeds on the fruit surface are protruding, the surface of the fruit is cracked and bronze color If the tree is slightly infected, the neighboring tree is not affected If the tree and ripe fruit are too heavily infected, the thrips will attack the neighboring plants and can spread throughout the strawberry orchard

Control measures:

Cultivation methods: Apply adequate and balanced fertilizers, prune old

leaves, collect and destroy residues

Chemical measures: Currently, there are no registered drugs in the list to

prevent this object It is possible to refer to the use of some drugs with active ingredients: Abamectin; Abamectin + Chlorfluazuron; + Abamectin + Emamectin benzoate; Abamectin 1.8% + Matrine 0.2%

2.6.2.2 Slug, viscous (Helix aspersa)

Morphological features and arising laws: The shell is thin, with 4 to 5

spirals, the color changes but is usually light chestnut gray, or brown with yellow streaks or spots The snail's body is soft and viscous, gray-brown, and collapses completely into the shell when not in operation When the slug pokes its head and legs out of the shell, the head has 2 pairs of tapioles The stubble can retract into the head Slugs are herbivores, eat at night, they eat a variety of plants

Figure 2.2 Helix aspersa Slug on strawberries

Harmful characteristics: Snails are often present in the field, but when it is sunny, they hide in shady and wet places such as dead leaves, nylon, rocks to spawn At night or on rainy days snails and slime crawl out to cause harm These lesions significantly reduce the value of the fruit and allow fungal pathogens to enter and grow

Control measures: Always keep the strawberry garden well-ventilated,

avoid high humidity in the field During the cultivation process, pruning leaves, collecting fruits if detecting slugs and slime must be collected Collect all bricks, stones in the field to limit the habitat of the slug species Use a plastic can mixed with substances such as beer baits or yogurt to trap the slug on the strawberry garden Currently, there is no registered pesticide to eliminate strawberry slug

2.6.2.3 White powder disease: (Sphaerotheca macularis)

Symptoms: Initially, the disease appears a white powder, which can be

seen behind the leaves, but on the leaves, stems, flowers, and fruits can also be infected Diseased leaves tend to curl upwards and reveal a layer of white powder behind the leaf surface Infected areas will usually wither and die

Figure 2.3 White powder disease on leaf

Infection process: Infected areas can disperse a large number of pathogens and follow the wind and spread through healthy plants

Fungi form on their own independent of moisture, and even under dry conditions fungi can be present This fungus is more common in greenhouses and plastic canopies than in outdoor farming White chalk fungus spreads very quickly and causes great losses to strawberry yield and fruit quality The disease damages the growth stages of the plant, but affects a lot in the flowering and fruit-bearing period

Figure 2.4 White chalk disease on strawberry plants

Control measures: Field sanitation, regular pruning of diseased leaf stems, and destroy them far away from the field Use disease-free seed sources Strawberry shading must be tall, ventilated, and high beds to avoid waterlogging in the rainy season Do not plant too thick density, increase potassium fertilizer for plants

2.6.2.4 Rubber disease: Fruit rot caused by Phytophthora cactorum

Symptoms: Both young and ripe fruit are discolored Green fruits are easy

to harden and turn brown Older fruits turn pale white, red or brownish and slightly soft The diseased fruit becomes dry, shrunk and chewy like rubber The special symptom of this disease is the loss of flavor and unpleasant odor Diseased fruit smells of car oil and has a bitter taste

Infection process: Fungi are present in the soil and enter the fruit due to the splashing of water when it rains or watering or because the fruit comes into contact with the planting soil The disease spreads and develops rapidly in the rainy season with high humidity

Figure 2.5 Rubber disease

Control measures: To control this disease, it is necessary to control the amount of water An inexpensive and effective way to prevent and prevent is to use a coating Hay and straw mulch is best But the mulch must be thick enough

that the fruit will not come into contact with the soil and will not be affected by water splashes

The best practice is to coat a layer of grass or dry straw on the surface of the soil and then cover the grass with a net This measure helps to keep the strawberry dry, not only prevents rubber disease but also prevents many other diseases

2.6.2.5 Black spot disease (Colletotrichum acutatum)

Symptoms: When the fruit is ripe, brown round spots appear Round spots

darken and then turn completely black If the fruit becomes infected before ripening, the whole fruit will become black and wilt During the process of transporting and storing, the disease continues to spread, making the fruit more damaged This is a matter of concern

Figure 2.6 Black spot disease

Infection process: The gooseberries are infected right from the planting time often do not show symptoms In mulberry fields, fungal pathogens can be spread by splashing water when watering or by heavy rain or through pruning and harvesting Plants with too much protein are also susceptible to disease

2.6.2.6 Gray mold: Fruit rot caused by Botrytis cinerea

Symptoms: Botrytis mushrooms mainly appear in the ripening stage, in humid conditions, the disease can cause serious harm Light brown spots appear first, then spread to the whole fruit and covered with a gray mold

Figure 2.7 Gray mold on strawberry plants

Young flowers and fruit can also become infected and cause the fruit to dry out The higher the temperature for storing the harvested berries, the faster the pathogen spreads

Infection process: Pathogens can come from infected leaves and fruits left

on the field and spread by the wind In addition, the pathogen can also come from outside the field, but it is not important Gray mold thrives in conditions of high air humidity and wet bed surfaces in rainy season conditions

Figure 2.8 Gray mold on strawberry diseases

Control measures: Clean up all residues of diseased plants that are burned

or buried away from the field Use a straw or net to prevent fruit from coming into contact with soil or moisture Choose a soil that is tall, well drained, and has

a high bed Fertilize NPK balance, increase potassium in the rainy season Rotation and soil treatment before planting

Limit the use of sprinkler systems, do not water at noon or late afternoon because this time to maintain the humidity will last Keep the surface of the strawberry bed always dry Harvested fruits should be stored at 2-40C to prevent fungal growth

Note: During times of high soil humidity and air humidity, it is necessary

to shorten the time between 2 sprays from 3-4 days of treatment to be able to limit the disease Spraying carefully on fruit clusters, when the flower blooms, avoid spraying with high concentration of spray and will cause deformity of the fruit In areas already resistant to drugs, different fungicides must be changed and used alternately Under prolonged and heavy rain conditions, fungal pathogens are difficult to control

2.6.2.7 Blackberry root rot disease

Harmful symptoms: Usually start within the first year of fruit

production Lesions will be most evident in lowland areas or areas with poor drainage The diseased tree grows poorly, lacks vitality, and is stunted Plants will experience water stress due to high water requirements during high growth, during or after flowering, or drought

On the leaves: At first, the leaves are red, like being boiled from the edge

of the leaves, then dry, then shake, making the plant wilt all the leaves

Figure 2.9 Blackberry root rot

The roots are blackened, and in the middle of the lybe of the central pillar, rot gradually spreads The diseased stem cuts the stem of the wood, turning yellowish brown at first, when the tree wilt and dies, the bruise spreads to the core and turns dark brown Plants with root rot disease often allow other fungal diseases to grow and damage stalks and fruits

Causes and development conditions of the disease: Black root rot is caused

by the complex interaction of fungi, nematodes and environmental factors Research by the Japanese Plant Protection Department from May 1973

to December 1974 showed that several fungi related to the disease include Rhizoctonia spp, Pythium spp and Fusarium spp When the roots are present with damaging nematodes, the disease is usually worse The disease usually occurs in lowland areas, easily submerged, on compacted soil or when organic matter is low

Usually one or more types of fungus attack are harmful Initial results of analysis, the harmful agents in Da Lat were fungi Pythium spp and Fusarium spp Environmental factors that are conducive to black root rot include poorly drained soils, low temperature damage, nutritional imbalances and herbicide effects, which become complicated and difficult to control

2.6.2.8 Red Spider: Tetranycus Urticae

Symptoms: attack on the underside of leaves, make young leaves turn yellow, dry due to depletion of nutrients, spider attacks on flowers, causing stamens to die ineffectively

Figure 2.10 Red Spider

Harmful characteristics: Both adults and larvae live concentrated on the underside of the leaf blades of young leaves that are gradually transitioning to the pudding stage

Spiders damage by injecting fluid of leaf tissue, causing the upper surface

of leaves to become yellow, patchy, brownish at the bottom You can see a small, fine spider web on the underside of the leaves If facing favorable conditions, spiders reproduce very quickly, the density can be up to several dozens per leaf, causing each leaf to become yellow and dry Flowers and fruits are also harmed by spiders Red spiders absorb nutrients in the fruit, making the fruit yellow, brown and cracked when the fruit is large Flowers can be shrunk, fall

Morphological characteristics and arising laws: Red spider has a very small body size, dotted with bran, and is difficult to detect with naked eyes adult spider 0.5-1mm long, pink, light red, spherical (female), the male is smaller, the body is oval, slightly pointed at the tail, the end ends are bright red, on the body and body has many hard bristles Maturity usually lasts 10-14 days

The adult lays eggs sporadically on the underside of the leaf blade Through the magnifying glass will see the eggs round, at first born pink white, then completely turn pink Eggs after laying about 4-5 days will hatch into young spiders The larvae are light green, when hatched only 6 legs, from the age of 2 onwards until they reach adulthood they have 8 legs The larval stage is about 6-9 days Each female lays 1 time from 50-100 eggs

Figure 2.11 Tetranycus Urticae red spider

Red spiders often arise and cause heavy harm during the hot dry season or during drought periods during the rainy season Red spiders spread by the wind,

by the fibers, the web they create

Prevention: Using natural enemies, insecticide Nissorun, Comite, Ortus, Oramíte,

Note: Applying the synthetic control measures to achieve higher results

than using single chemical methods Only use pesticides that are on the List of pesticides permitted to be produced, traded and used in Vietnam

2.7 Nutritional value and economic value

Strawberry Nutritional Value for many centuries before, strawberries had been a favorite among the fruits of the temperate world They were valued for delicious flavor and fragrance, for health-restoring qualities and as harbinger of spring (Wilhelm, S and J E Sagen., 1974)

Flavourful and nutritious, strawberries are enjoyed by millions of people in all climates, including and are predominantly used as fresh fruit Their use in processed forms such as cooked and sweetened preserves, jams or jellies and

frozen whole berries or sweetened juice extracts or flavorings, and their use in making a variety of other processed products made them one of the most popular berry crops, more widely distributed than any other fruit (Samir et al., 2007)

Of its many positive characteristics, the nutritional value of strawberries is nearly perfect Eight medium strawberries contain more Vitamin C than an orange, 20% of the recommended daily allowance for folic acid, no fat, no cholesterol and are considered high in fiber (Driscollís 2004)

Strawberry is cultivated all around the world, not only for its digestive and tonic properties, but because of the nutritional value of its fruits, important source of folate, Vitamin C, fiber, potassium, flavonoids, autocianidin, phytochemicals and antioxidants

Strawberry Phytochemicals and Human Health

Although strawberry is not an essential component of the diet, its delicious flavor and taste, attractive appearance and seasonal availability make this fruit

an excellent crop Even more, strawberries are rich in phytochemical compounds with potential antioxidant compounds, mainly ellagic acid and flavonoids, which can lower the risk of cardiovascular events and tumorogenesis (Hannum, S.M., 2004)

These qualities have ensured that the economic importance of this crop has increased throughout the world and, nowadays, it remains as a crop of primary interest for both research and fruit production (Mercado et al., 2007)

A growing body of data suggests that consumption of a phytochemical-rich diet reduces the risk of certain chronic human illnesses such as cancer, heart and neurodegenerative diseases Strawberry (Fragaria x ananassa Duch), fruits are a rich source of phytochemicals (plant chemicals) of which phenolic compounds predominate Berry fruits are reported to contain a wide variety of phenolics including hydroxybenzoic and hydroxycinnamic acid derivatives, anthocyanins, flavonols, flavanols, condensed tannins (proanthocyanidins) and hydrolyzable tannins

Studies conducted in vitro indicate that berry phenolics have a wide range

of biological properties such as anti-cancer, antioxidant, anti-inflammatory, and cell regulatory effects (Seeram, N.P., 2006)

2.8 Factors regulating development

There are two primary types of strawberries now grown commercially, neutral and short day plants Long day ("everbearing") plants are also available, but they are rarely grown outside of home gardens The short-day types are actually facultative short-day plants and initiate flower buds either under short day conditions (less than 14 hrs of day length) or when temperatures are less than 15°C (Darrow, 1936; Guttridge, 1985; Larson, 1994) Above 15°C, the critical photoperiod for floral induction is between 8-14 hours, depending on the cultivar

day-In mild tropical climates, the short-day cultivars developed for temperate climates can be highly productive (Subramanium and Iyer, 1974); although flower bud formation is restricted if temperatures get too hot (Strik, 1985) Durner et al (1984) observed no floral induction under short days in plants of F

x ananassa held at 26/22 and 30/26°C day/night temperature regimes Chabot (1978) found that no flowering occurred in the related species, F vesca, above a 20/1O°C temperature regime In tropical highlands with short days and cool climates, short-day cultivars of F x ananassa perform similar to day-neutrals in temperate climates, with cyclical patterns of flowering and fruiting

The minimum number of photoinductive cycles necessary to induce flowering in short day plants has been variously reported to be 7-24, depending

in part on temperature (Hartmann, 1947 and b; Went, 1957; Guttridge, 1985; Larson, 1994) At higher temperatures, longer photoinduction periods appear to

be needed Ito and Saito (1962) found that under 8-hr photoperiods only 10 cycles were necessary for floral induction at 24°C, while at 30°C more than 20 were required With 16-hr photoperiods, 10 cycles were necessary at 9°C, but 16 were required at 17°C

While short-day cultivars adapted to cooler climates can be grown in tropical and sub-tropical regions, they have a tendency to require a chilling period for full productiv- ity Those cultivars developed for warm regions may not need one The ability to grow well during the short days of October, November and December in North American greenhouses has classically been used as an indicator of a cultivar's regional adaptation Those adapted to cooler regions generally grow poorly during this period and enter a rest period, while those adapted to warmer climates continue to grow (Darrow and Waldo, 1934)

A new type of short-day strawberry has recently been developed in Israel for tropical and sub-tropical environments (Izsak and Izhar, 1984; Izhar and Izsak, 1995) These "infra" short-day types do not require chilling for high yields and initiate flower bud primordia in response to longer light regimes (13.5

to 14 hrs.) and higher temperatures (1O-26°C) than traditional short-day plants These genotypes fill an important economic window as they are productive in the fall and early winter when short-day varieties are incapable of bearing fruit Day-neutral plants have been developed for temperate climates that produce crowns and flower buds approximately 3 months after planting, regardless of the day length (Bringhurst and Voth, 1975; Galletta and Bringhurst, 1990) They have the potential to initiate flower buds cyclically throughout the growing season, although high temperatures appear to inhibit bud formation Durner et al (1984) found that day/night temperatures of 30/26°C almost completely inhibited flower bud initiation in the day-neutrals 'Hecker' and 'Tristar', while these cultivars were highly productive under 18/14, and 22/18°C regimes

While most cultivars are now categorized as day-neutrals or short-day plants, some genotypes are hard to rank precisely due to complex interactions between genotype, temperature and photoperiod (Nicoll and Galletta, 1987; Yanagi and Oda, 1989) Darrow (1966) suggested that strawberries actually range from obligate short day to facultative short-day to complete day-neutrals The infra short-day types developed by Izsak and Izhar (1984) fall in the middle

of this range Darrow also suggested that a range of everbearers types exist from weak, intermediate to strong Galletta and Bringhurst (1990) have suggested that flowering in modern strawberry cultivars is regulated more by sensitivity to high temperature than photoperiod, with traditional short-day plants being more sensitive to high temperatures than day-neutrals

2.9 Research in the world and in Vietnam

Benzyladinine seems to be important in controlling proliferation of strawberry shoots Marcotrigiano et al (1984) observed on a large number of strawberry cultivars that BA concentrations in the range 1.3-13.3 mM, did not produce significant differences in multiplication rate; in contrast, Simpson et al (1989) found clear differences among genotypes in their BA requirements for optimum shoot proliferation

Singh and Pandey (2004) reported that half strength MS medium supplemented with 1 mg / l BA and 1 mg/l IBA produced the highest number of shoots (13 per explant)

Kaushal et al (2006) found that callus of strawberry differentiated into shoots after transferring to MS medium supplemented with 0.5 Kin + 2 mg/l BA + 0.25 mg/l NAA Well developed shoots were transferred to multiplication medium containing 0.5 mg/l Kin + 0.5 mg/l BA and 1.0 mg/l GA3

Weifeng et al (2004) revealed that Kin used together with BA for adventitious bud induction gave better results than Kin alone, but, Waithaka et al (1980) revealed that axillary dormancy was released by including a large level (50 µM) of Kin in the culture medium

In that concern, some investigations did not recommend full MS for strawberry rooting, Kaushal et al (2006) found that rooting of strawberry was done in MS half strength with 1.0 mg/l IBA and 0.2 mg/l activated charcoal Gautam et al (2001) indicated that the highest root induction frequency obtained was 95.23% on 1/4 MS medium with IBA at 1.0 mg/l and charcoal (200 mg/l)