If you have the belief that you can do it, you can surely acquire the capacity to do it even if you don t have it at the Beginning

Transcription

1 INTRODUCTION If you have the belief that you can do it, you can surely acquire the capacity to do it even if you don t have it at the Beginning

2 Chapter 1 INTRODUCTION Flowers have been associated with man's history from Neolithic age and being the part of human life from birth to death. Man has traditionally used flowers for expressing or exhibiting his innermost feelings to God and deities or presenting it to beloved ones or complementing anyone or versifying any conceivable emotion. Thus, the importance of flowers in socio-cultural and religious life of the humans can hardly be exaggerated. Floriculture which is an ancient farm activity with immense potential is now not only generating remunerative self-employment among small and marginal farmers but also earning the much needed foreign exchange. With the urban Indians becoming more westernized, floriculture has become an exceedingly lucrative business now-a-days. Presenting bouquets on birthdays, weddings, anniversaries, for wishing good luck or get well soon and other occasions, sending flowers as seasonal or festival gifts is becoming common practice (Arora, 2008). All over the world, the floricultural sector is experiencing rapid changes. Due to globalization and its effect on income generation in different parts of the world, the per capita consumption of flowers in most countries is increasing. Besides traditional centers of production viz. The Netherlands, Columbia, Israel and Kenya, new production centers are developing in Latin America, Africa and Asia and the production has increased many times, as compared to what it was a few decades ago. The Asian countries like India, China, Korea, Vietnam, etc., are moving in the direction of more intensive floriculture (Kapoor, 2001). Commercial floriculture has immense significance in the diversification of agriculture and in national economy. The area under flower crops in India has risen to approximately 1.91 lakh hectares with an estimated production of million cut flower stems and lakh metric tons of loose flowers with productivity of 5.4mt/ha. There is an export of metric tons of flowers, valued ` million from India (NHB, 2011). With the global boom in floriculture trade, floriculture in India has come up as one of the primary thrust areas for procurement of export quality flowers. Indication of global deficit of flowers, paints a rosy picture for India. Impressed by the prospects of the growing market, the government has formulated major action plans for floriculture. The growth of floriculture industry has taken long strides worldwide, especially in the developing countries as a result of outsourcing which is due to low cost of maintenance

3 2 including low labour costs. The important flower crops in the international flower trade are rose, carnation, gerbera, chrysanthemum, gladiolus, orchids, lilium etc. The demand of cut flower is increasing day by day with increasing standard of living, aesthetic sense and awareness in the people. Chrysanthemum belongs to family Asteraceae or Compositae (subtribe Chrysantheminae, tribe Anthemideae) also called the aster, daisy, or sunflower family. The name Asteraceae is derived from the type genus Aster, which is a Greek term, meaning "star" ( while Compositae, an older but still valid name (ICBN Art. 18.5, 2007). It has approximately 1,620 genera and more than 23,600 species, is the largest family of flowering plants (Stevens, 2001). In India, 900 species in 167 genera (Hajra et al. 1995) are reported. The family is further subdivided into 12 subfamilies (Tree of Life - Asteraceae), dominated by the large subfamily Asteroideae that contains more than 70% of the species of the family. The tribe Anthemideae is one of the largest tribes of the family with 1,741 species predominantly distributed in Eurasia, North and South Africa, with fewer species in North America and Australia (Bremer and Humphries, 1993). Chrysanthemum is a popular commercial flower grown for cut flower as well as pot plant (pot mum). It is an important global floriculture crop leader due to its unparalleled diversity in plant and flower colour, shape, form and blooming pattern. In international florists trade, chrysanthemum ranks third as a cut flower and fifth as pot plant (Anonymous 2008). The genus Chrysanthemum consists of more than 40 species distributed mainly in East Asia. More than 20 species are believed to be a native of China where it has been cultivated for more than 1600 years (Zhao et al., 2009). In India, the first written mention of chrysanthemum is in Marathi literature in Gyaneshwari (1290 AD, Chapter 15, Shaloka 20), a treatise on Geeta by Sant Gyaneshwar (Datta and Bhattacharjee, 2001). In 1960, H van Rheede reported a cultivar from India called Gool-doodi grown in Holland (Kher, 1988). The most evident characteristic of Asteraceae is perhaps its inflorescence: a specialized capitulum, technically called a calathid or calathidium, but generally referred to as flower head or, alternatively, simply capitulum (Usher, 1966). The unique inflorescence involucrate capitulum, which is densely packed cluster of numerous small individual flowers called florets. The outer perimeter of flower head possessing a long strap like petal

4 3 termed as ligule (ray florets). The inner portion of the flower head is composed of small florets (disc florets), composed of tubular corollas. The ray florets are pistillate, while disc florets are fertile and perfect (Dwivedi and Banerji, 2011). The members of family are characterized by the capitulum inflorescence, calyx replaced by pappus, syngenecious condition of anthers, bicarpellary, syncarpous inferior ovary with single basal anatropous ovule and achenaceous fruit. Inspite of these common features the members of the family show lot of variation with respect to capitulum characters. Different tribes, genera and species are demarcated on the basis of capitula characters (Burtt, 1977). The genus Chrysanthemum is characterized by annual or perennial herb or woody nature; leaves alternate dissected, terminating long peduncles in cormybose clusters, involucre scales are imbricated and appressed. Now, the genus has been split into many genera as Argyanthema, Chrysanthemum, Leucanthemum, Tanacetum, Dedranthema, etc. Among the ornamental plants of family Asteraceae, Chrysanthemum is one of the widely cultivated and popular composit due to the immense variation in number and diversity in capitula shape, size and colour displayed by it contribute to its popularity in the cut flower industry. Genus Chrysanthemum has a lot of cytological variability with a basic chromosome number of nine (x=9), however a ploidy level ranges from 2n=18 to 45, 47, (Nazeer and Khoshoo, 1982). Polyploidy has played an important role in chromosome evolution in this genus from diploid to decaploid (Abd El-Twab and Kondo, 2003 and Abd El-Twab et al., 2012). Many perennial species exhibit a higher degree of interspecific polyploidy (2x to 25x), whereas the annuals are strictly diploid and have exploited reciprocal translocations for adaptation and speciation. The evolution in the genus has occurred mainly due to polyploidy, aneuploidy, structural changes, hybridization, apomixis, etc. The advent of molecular biology has influenced the way in which people think about creating useful variability. Molecular biology offers powerful analytical tools for analyzing genetic alterations, including induced mutations. For genetic diversity analysis and cultivar identification, the DNA markers have many advantages over the traditional morpho-biochemical markers and may provide desired momentum to accelerate the pace of crop improvement programme. Among the DNA markers, polymerase chain reaction (PCR) based markers using arbitrary primers, such as random amplified polymorphic DNA

5 4 (RAPD) have been widely used for investigating genetic relatedness and diversity in population and cultivars (Li and Nelson, 2001). Around 1990, the polymerase chain reaction technology became available (Erlich et al., 1991). This technique enabled one to show differences at the DNA sequence level in an easy and fast way, even if only small amount of tissue are available. William et al. (1990) developed the use of general purpose decamer primers in the RAPD technique. With these single primers arbitrary fragments were amplified and generally high level of polymorphism was obtained, without any prior knowledge of the DNA sequence amplified. Since then, chrysanthemum cultivars and other closely related family members of Asteraceae have been characterized based on RAPD (Wolff, 1996 and Martin et al., 2002). Wolff and Rijn (1993) showed that with the RAPD technique reproducible patterns were obtained in chrysanthemum and that with these patterns cultivars could be distinguished. It was, however, also demonstrated that cultivars from one family, differing for flower colour, could not be distinguished with several DNA techniques (Wolff et al., 1995). Genetic variation between two genetically diverse tissues and chimera cell types in leaf has been demonstrated using the RAPD technique (Wolff, 1996), but this is limited only up to the characterization of the different varieties of Chrysanthemum morifolium. Identification and characterization of plants is extremely important in horticultural crops in order to protect the Plant Breeders Rights. Earlier new varieties were identified based on horticultural and physiological parameters (Kumar et al., 2006). The main advantage of RAPD technique lies in its rapidity, applicability to any organism without prior knowledge of nucleotide sequence and its ability to detect mutations. The breeding of ornamental plants started centuries ago but, reliable data are only from the last years. Within past 80 years, progress in producing new types and forms has accelerated largely due to the need for higher yields, quality and disease resistance (Misra et al., 2006). The quest for developing new varieties in flower crop is never ending. There is always a craze for developing new varieties by replacing older varieties with newer ones. Since, in flowers a specimen cannot maintain interest for a long time; people have the desire to develop newer forms through various methods of breeding. The interest has been generated to evolve new forms, new colours and their large scale production. Interest in flowers among floriculturists and commercial growers immensely increased due to its blooming export market since the last few decades.

6 5 Basically, the aim of the breeder is to produce improved plants but the kind of improvement is very wide, from the visible attributes of flower colour, form and size, plant height, leaf colour, presentation and markings, to the less obvious but equally important disease resistance, cold hardiness, flowering date, forcibility and the keeping quality of the cut flowers as well as some less important aspects like scent. For a new line to be successful; it must possess large number of desirable characteristics; failure in one major area means rejection, although it might still have value in a breeding programme to preserve its good qualities. The primary objective of plant breeding programme is to create variability and to select the best recombinants possessing desirable characteristics. Selection is effective only in the presence of considerable variability in population. Therefore, genetic variability is the prerequisite for an effective breeding programme in many crops and it differs from crop to crop due to its evolutionary process and complexity of hereditary system. Induced mutations could be one of the approaches for the creation of genetic variability. The possibilities of mutation breeding in vegetatively propagated species are favourable in general for various reasons such as the usually large heterozygosity of the material which allows direct detection of mutations in the irradiated material. In the vegetatively propagated material in which the intention is often improvement in visible characteristics, selection of potentially useful mutations is generally easy (Broertjes, 1968). The industry must meet the demand of consumers by providing new value added flowers which are cost effective and unique in flower characteristics e.g., flower shape, flower colour and have long shelf life. Mutation assisted breeding (MAB) together with biotechnology can contribute greatly for genetic improvement of ornamental plants and in uplifting the socio-economic benefits in the developing countries (Jain, 2006). Mutagenesis is a unique force in creating variation as it may alter even those genes that are common to all the varieties of a species and, therefore, do not show any segregation after a cross within that species. Favourable induced changes can be selected and also be incorporated, through crossing and selection in a new improved genotype. The chrysanthemum shows high heterozygosity, strong sporophytic selfincompatibility as shown by all the members of Asteraceae family (Richards, 1986). The chief mechanism in cytological evolution and diversity of chrysanthemum are out-

7 6 breeding, spontaneous and intentional hybridization, coupled with mutation, chromosomal differentiation and re-patterning and polyploidy. The modern day cultivars of C. morifolium are resultant of various breeding techniques like selection, crossing, mutation and biotechnological tools. About 1 / rd 3 of the commercial chrysanthemum varieties have originated as sports or somatic mutations (Maluszynski et al., 2000). Mutation breeding has been recognized as a useful complimentary tool for the improvement of modern day chrysanthemum and also source of creation of variability. Induced mutations are highly effective in enhancing natural genetic resources and have successfully assisted in developing improved and new cultivars. Among more than 2300 officially released mutant varieties worldwide, 566 represent ornamental plants (Jain, 2006). Chrysanthemum appears to be ideal system for mutation breeding as many characters of economic importance are easily monitored after mutagenic treatment, without negative, residual effect on human health. It is known that frequency and spectrum of mutations differ somewhat depending upon the mutagen used and the dose applied. The physical mutagens are widely used and have the advantage of good penetration and precise dosimetry (Micke and Donini, 1973). The difference in the genetic background behaved differently for a mutagen. Genetic differences even though they are as small as a simple gene difference, can induce significant changes in the radio-sensitivity which influences not only the total rate but also the spectrum of recoverable mutations (Gustafsson, 1944; 1947 and 1965). Induced mutations consequently have a high potential for bringing about further genetic improvement. Induced mutations are already widely used in commercial breeding of ornamentals plants. The genetic variability can be induced by mutagenic agents, such as radiations and chemicals, from which desired mutants could be selected (Jain and Spencer, 2006). The mutagen treatment breaks the nuclear DNA and during the process of DNA repair mechanism, new heritable mutations are induced randomly. The changes can also occur in cytoplasmic organelles, and also result in chromosomal or genomic variations that enable plant breeders to select useful mutants such as flower colour, shape, disease resistance, early flowering types (Jain and Maluszynski, 2004). In vegetatively propagated plants, the advantage of induced mutation is the ability to change one or few characters of an outstanding cultivar without altering the remaining genotype and any

8 7 induced colour sports can be used directly for propagation as a cutting without further breeding (Broertjes, 1972). Physical and chemical mutagens offer possibility for the improvement of chrysanthemum cultivars. Ionizing radiations viz. X-rays, γ-rays, high ion beam (HIB) have been reported to be the best mutagens for induction of somatic mutations in chrysanthemum. Among these γ-rays irradiation has been commercially used effectively for inducing mutations in chrysanthemum with dose ranging from 1 kr to 10 kr. More mutations have been documented in the pink cultivars as compared to others (Datta and Banerji, 1993). Acute radiation and chronic radiation are two ways to irradiate with gamma radiation. In case of acute radiation high dose is given for a short period of time (seconds, minutes and hours) whereas in case of chronic radiation low radiation dose treatment is given for a long period (days, weeks, months and years). Chemical mutagens like EMS (ethyl methane sulphonate), MMS (methyl methane sulphonate), DES (diethyl sulphate), etc. are often applied for induction of somatic mutations in chrysanthemum. Scientific interest in mutation breeding has drastically decreased during the last two decades. Interest in research has shifted towards development and application of molecular techniques as tools in breeding and transformation of plants, since they allow more directed approach in pursuance of breeding goal. Molecular techniques generate high developmental costs and require sophisticated equipments and a highly trained staff. The investment in such expensive methods does not seem adequate in case of many ornamental species with their limited economic importance as compared to agricultural crops. Genetically modified ornamental plants are not marketed in Europe due to low acceptance of consumers and the uncertain legal situation (Schum, 2003). The breeding of a photo-insensitive variety chrysanthemum cultivar Horim through induced mutation in the Netherlands brought revolution during 1979 in the international chrysanthemum trade by registering a record sale of more than 150 million cuttings worth 95 million US$ (Broertjes et al., 1980). Recently the interest in evolving novel mutants is growing with the possibility of getting new cultivar and patenting it. The main advantage of mutagenesis in chrysanthemum is the possibility to alter one or few characters of the outstanding cultivar, without changing the rest of the genotype, thus saving time and money for breeding a new cultivar.

9 8 Objectives Keeping above possibilities in view the present investigations on Morphological, physiological, biochemical, molecular and mutational studies in different species of Chrysanthemum were carried out with the following objectives: 1. Morphological and physiological characterization of the different species of Chrysanthemum. 2. To characterize the different species of Chrysanthemum by using biochemical and molecular techniques to supplement morphological descriptors. 3. To screen out the mutants for ornamental traits, thus developed as a result of mutagen treatments on different species of Chrysanthemum.