I INTRODUCTION

Areca nut (Areca catechu L.) is the most profitable plantation crop grown in humid
tropics of India, realizing highest economic returns per unit area. Apart from tobacco, alcohol
and caffeine, it is the most commonly used substance and globally, an estimated 600 million
people are believed to indulge in the habit (Raja et al. 2007). Areca nut is usually consumed
alone, in its natural form, as a processed product,or as a key ingredient of the chewing quid
(Warnakulasuriya2002). The exact origin of areca nut palm is not fullyknown as there are no
fossil records of the genus Areca.Geologically the palm extends to the Cretaceous period
ofupper Mesozoic era (Mahabale 1982). It is a native ofCochin (China), the Malaya Peninsula
and neighbouringislands (Watt 1889). The exact native country of areca nutis uncertain (de
Condolle 1886) but the range of distributionin a locality where the plants are thought to be wild
maythrow some light on its origin (Furtado 1933). The historyof areca nut chewing was
established among the Aryans atleast 2000 years ago (Gode 1961).

Areca nut plays a prominent role in the religious, social,cultural and economic life of
Indians. The economicproduct is the fruit called “betel nut” and is used mainly formasticatory
purposes. In India, areca nut is used as an offeringduring religious ceremonies (Murthy 1968).
It is cultivatedin the plains and foot hills of Western Ghats andNorth Eastern regions of India.
Area and production indifferent states show that Karnataka, Kerala and Assamaccount for over
90% (Rajagopal 2004). It is vital to smallland holders as a source of sustainable income. It is
grownover an area of 2,64,000 ha with an annual production of313,000 t (Chowdappaet al.
2003). It is estimated thatnearly ten million people depend on the Areca nut industryfor their
livelihood in India. The quality, variety and typesof Areca nut vary from one place to
another.According to Bentham and Hooker, the genus Areca isthe first one in the family
Palmae, in their treatise SpeciesPlantarum. The genus expanded rapidly from its
monospecificstatus and it is believed to compose of about 76 species.Among these, A. catechu
is the only cultivated species andnuts of a few other species such as A. triandraRoxb. and
A.concinnaThw. are also used as a masticatory (Murthy andPillai 1982).

Chewing areca nut is the most common modeof administration although its use as a
masticatory has beenon the decline over the passed few decades, with the incursionof modern
ways of living among rural populations,particularly in South East Asia (Rajagopal 2004).
Recentclinical diagnostic studies have revealed that chronic arecanut chewing has been
strongly linked to the development oforal submucous fibrosis (Murtiet al. 1995;
Warnakulasuriyaet al. 1997; Gupta et al. 1998), a chronic progressivepre-cancerous condition
characterised by fibrosis of themucosal lining of the oral cavity and occasionally the
upperdigestive tract (Maher et al. 1991). The development ofcancer in the upper aero-digestive
tract is increased markedlyin patients with consumption of areca nut (Warnakulasuriya2002).

Diversity of areca nut germplasm based on morphological characteristics has been

performed. Beccari (1919)recognised four cultivars of areca nut and termed them asvarieties
silvatica, batanensis, longicarpaand communisbased on the size and shape of fruit and kernel.
Based ontheir height areca palm are classified into tall, semi-tall anddwarf types. The height
ranges between 60 and 360 cm asrecorded at the seventh year of age (Rajagopal 2004). Murthy
and Bavappa (1962) identified 64 cultivars based onfruit size, from Kerala, Karnataka and
Maharastra. Based onthe variation in stomatal characters, concerning number ofstomata per
unit area four cultivars were identified by Bavappa (1966). Bavappa and Pillai (1976)
foundhighly significantdifferences in respect of number of leaves shed,spadices and female
flowers produced, nut set, number ofnuts harvested and, weight and size of nuts among

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.1

 thirteencultivars of A. catechu from eight countries. Cultivars availablein Malaya, Sri Lanka
and South India have been designatedby local names (Aiyer 1966; Nambier
1954).Identification of cultivar and estimation of geneticdiversity using phenotypic markers
have several limitations,especially in perennial crops. Molecular differences, whichcan be
detected using DNA and protein-based markers, aremore authentic and unaffected by
environmental factors(Dhanraj et al. 2002).

Hence, the characterisation of genotypesat the genetic level provides the first step
towardsmore efficient conservation, maintenance and utilisation ofexisting genetic diversity
(Prakash et al. 2002). AmongDNA-based molecular markers, SSRs (Simple sequence repeats)
proved as excellent tool to estimategenetic diversity and relationships among genotypes (Hu et
al.,2011). They are simple, versatile, relatively inexpensive,and can detect slight genetic
differences andhelp in identifying duplicates in the populations. SSR markers also have been
used successfully to study geneticdiversity and relatedness among perennial crops by our
researchgroup such as mango (Hemanth Kumar et al. 2001),guava (Prakash et al. 2002),
cashew (Dhanraj et al. 2002)and jackfruit (Simon et al. 2007). To the best of our knowledgeno
research work on genetic studies has been carriedout on areca nut cultivars despite its
importance. Hence, the present study has been planned with the following objectives.

1. To analyse the genetic diversity/ genetic relationship among arecanut germplasm/

accessions using simple sequence repeat (SSR) markers.
2. To study the genetic diversity and variability of various morphological characters in
arecanut germplasm.
3. To estimate the arecoline content in arecanutgermplasm.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 II REVIEW OF LITERATURE

The conservation of wild species, local varieties and traditional genotypes in seed gene bank
has become a useful tool of gene maintenance. The accessions in gene banks should be characterized
and evaluated in order to determine the magnitude of genetic diversity which would allow the
identification of redundant accessions and genotypes of interest in breeding program. Only a few
previous reports are accessible now for its research which can lead to uprising in evaluation of
arecanut biology. Few studies have been done using different genes to examine its genetic variability.
Literature available concerning to the present study has been presented below.

2.1 Origin and domestication of arecanut

The palm Areca catechuL., Arecaceae is known variously as arecanut ,betel nut and adike. The
center of arecanut domestication likely lies in eastern Asia, possibly eastern India or southern china
(Minirajet al., 1993; Walters and Decker Walters, 1988). Uncarbonized seed coat fragments have been
tentatively identified from spirit cave in northern Thailand. However there have been no
archaeological reports of arecanut remains in China (Marr et al., 2004). Moreover, a comprehensive
compilation of plant remains from 124 Indian archaeological sites does not include arecanut (Kajale,
1991). Wild or small nuted cultivated forms however are mentioned in Ayurvedic texts written in
Indian Sanskrit from 2000 to 200 BCE by members of the Indian Aryan culture (Decker Walters,
1999). Both the domesticated and putative wild arecanut progenitors of arecanut are listed in floras of
India, 5 tropical Africa and Asia as well as the new world tropics, where it first arrived in Brazil via
the slave trade from Africa and then spread into central America (Marr et al. 2004). Based on both
historical literature (Minirajet al., 1993, Chakravarty, 1990, Walters and Decker Walters, 1988) and
recent random amplified polymorphic DNA (SSR) (Dey et al., 2006), inter simple sequnce repeats
(ISSR) (Singh et al., 2007) and amplified fragments length polymorphism (AFLP) (Gaikwad et al.,
2008) molecular analysis, western ghats of India may be considered as a probably primary center of
diversity of arecanut, where a number of wild species (Chakravarty, 1990) currently exist.

2.2 Variability and genetic parameter

The assessment of variability present in any crop species is an essential prerequisite for
formulating an effective breeding program, as the existing variability can be used to enhance the yield
level of cultivars following appropriate breeding strategies (Patil et al., 2012). Sreelathakumary and
Resmi (2015) observed that the characters of 33 arecanut genotype showed ample variation as the wide
range obtained for days to seedling emergence, palm length and internodal length. Days to first male
and female flower plays an important role in deciding the earliness or lateness of the crop. The early
and late female flower appearance helps in occurrence of early or late flush of the crop. Days to first
male flower showed wide range of variations among the genotypes.

Parameters of genotypic phenotypic coefficients of variation (GCV and PCV) are useful in
detecting the amount of variability present in the available genotypes. High phenotypic co-efficient of
variation than the genotypic co-efficient of variation indicates more influence of environment on the
expression of genes controlling the trait. Therefore, it can be referred that selection based upon
phenotypic expression of the particular character wouldn’t be productive for the improvement of that
crop (Bhuiyan, 2014). If the genotypic co-efficient of 6 variation and phenotypic co-efficient of
variation are close to each other, it suggests environmental influence is minor on the expression of the
genes controlling the trait. So, selection based upon phenotypic expression of that character would be
effective for the improvement of the specific crop.

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.3

 Rahman et al. (1991); reported that male flower were earlier than female flower in several
genotypes of bottle gourd, ribbed gourd and sweet gourd. They reported significant variations for that
character among the genotypes of arecanut, sweet gourd, ribbed gourd and bottle gourd. Significant
variation for it length and diameter were also observed. Abusaleha and Dutta (1990), carried out a
study with 65 genetic stocks to assess the genetic variation and heritability in ridge gourd. Significant
variability was observed for all the characters at phenotypic as well as genotypic level with a very
wide range of values. It was observed by Rahman et al. (1990) in a study that significant variation for
days to first flowering among the genotypes of arecanut, ribbed gourd and sweet gourd. Rahman et al.
(1990, 1991); also concluded that days to flowering was earlier than days to female flowering in
several genotypes of ribbed gourd, arecanut, bottle gourd and sweet gourd. They also reported that
arecanut,

2.3Molecularcharacterization

Puroshothamet al. (2008) conducted a study on Genetic relationship between cultivar of Areca
nut (Areca catechu L.) determined by RAPD. The experiment was conducted at plant biotechnology
laboratory, UAS, Bengaluru. Eleven cultivars of Areca nut (Areca catechu L.) obtained from Western
Ghats regions of India were analysed using RAPD markers. PCR-amplifiable DNA was isolated using
the CTAB method and 14 amplified fragments were obtained using 19 random primers. The genetic
dissimilarity matrix, which was calculated based on Square Euclidian Distances revealed a maximum
genetic distance of 47% between cultivars, ‘Mohit Nagar Interse’ and ‘Mohit Nagar’ and the minimum
genetic distance (22%) was observed between the genotypes ‘Maidan Local’ and ‘Sree Mangala’. The
Ward’s method of cluster analysis grouped all the individuals on a dendrogram into two major clusters
‘A’ and ‘B’ twentynine linkage distances with two sub-clusters in cluster ‘A’. The sub cluster ‘A1’
consisted of nine cultivars in two minor clusters ‘A1a’ and ‘A1b’ linked at tentyseven distances. The
sub-cluster ‘A2’ consisted of one cultivar ‘Sumangala’ linked with ‘A1’ at 28 linkage distance. The
cluster ‘B’ consisted of one cultivar ‘Mohit Nagar’. The study showed moderate genetic diversity
among the cultivars of Areca nut. The RAPD analysis proved to be a quick, simple and significant
testing method to assess genetic diversity among Areca nut populations studied.

Bharath et al. (2015) conducted a study on Genetic relationship and diversity in Areca nut
(Areca catechu L.) germplasm utilizing RAPD markers. The experiment was conducted at ICAR-
CPCRI Regional Research Station ,Vittal. Sixty different areca nut accessions were selected from the
140 accessions which are conserved and maintained in the field gene bank of CPCRI ,Vittal.From each
accession leaf samples were collected from four different palms representing an accession. DNA was
extracted using standardized protocol (Rajesh et al., 2007) . RAPD analysis was done after screening
areca nut accessions with random decamer Primers of the series OPM, OPAF AND OPC to identify
polymorphic markres. DNA band size was estimated by comparing the DNA bands with a 1 Kb DNA
ladder as reference The binary data matrices of before 60 different accessions were analyzed using
NTSYS pc Package to generate Jaccard’s similarity coefficient. The similarity co-efficients were used
to construct dendrogram using UPGMA and SAHN routine. The same binary data was also subjected
to principal component analysis (PCA) with NTSYS software package. Maximum similarity was
observed between Shriwardhana-M and Shriwardhana-B(0.93) and the minimum similarity was
observed between Diveagar-I and Saigon-I(0.68).The Dendrogram revealed two major clusters which
appear to be equally divergent at 68 percent level.Two accessions having least similarity Diveagar-I
and Saigon-I could be used for hybridization studies in order to obtain highly heterotic progenies.

Hu et al. (2009) described the development of nine microsatellite loci from A. catechu. The
number of alleles per locus ranged from 5 to 15. The expected and observed heterozygosities ranged
from 0.71to 0.94 and from 0 to 0.88, respectively. All microsatellite loci, except for AC30,

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

significantly deviated from Hardy–Weinberg equilibrium possibly due to artificially selected
cultivation or the existence of excessive null alleles. No linkage disequilibrium was observed from
pairwise comparisons of loci, except for AC06 and AC08.

2.4Morphological characterization

Niralet al. (2013) conducted a study on Morphological and molecular characterization of a

large fruited unique coconut accession from Vaibhavwadi, Maharashtra, India. The experiment was
conducted at Central Plantation Crops Research Institute (CPCRI), Kasaragod. Forty three accessions
from around the world. Twenty three morphological traits, six tender nut traits and eight fruit
component traits were used for the study of Morphological variability. DNA was extracted from the
leaf samples of each accession through the column purification method of the Himedia plant genomic
DNA extraction kit. Fruit component analysis of the nuts of this accession indicated that they have low
husk to nut weight ratio typical of the Niuvaitype cultivated in South East Asia. The microsatellite
analysis indicated that Vaibhavwadi coconut population (MAHT) is generally close to the South East
Asian coconut accessions and has proximity with dwarf accessions in the conserved germplasm of
India. The probable origin of this type was identified as Borneo Tall (60% similarity) a cultivar which
is known to produce large coconuts.

Samsudeenet al. (2013) conducted a study on In situ approach for rapid characterization to aid
on farm conservation of coconut germplasm. . The experiment was conducted at Central Plantation
Crops Research Institute (CPCRI), Kasaragod. Identification and location of desirable ecotypes in its
natural home was done through participatory assessment tools like key informant interviews and focus
group discussion. A total of nine attributes were considered for this purpose. Two genotypes Bedakam
and Kuttiyadi were selected and ten each of morphological and reproductive characters along with
fourteen fruit characters were recorded. Comparison of the two ecotypes revealed that the traits, trunk
girth, length of internode, number of leaves, number of bunches with nuts, number of nuts, shell
weight, husked fruit weight and fruit weight were higher in Kuttiyadi than in Bedakam ecotype. On the
other hand, number of leaf scars per meter, length of inflorescence, fruit breadth, husk weight, nut
cavity volume and copra weight were higher in Bedakam compared to Kuttiyadi ecotype.

2.5. Estimation of arecoline.

Sankaran Nambudiri(1968) conducted a study on estimation of Arecoline. The experiment was

conducted at Central Food Technological Research Institute, Mysuru. Nuts from the Areca nut palms
were collected, dried and powdered. To standardize the conditions for estimation, the parameters like
particle size of the dried areca nut powder, number of extractions, Alkali concentrations, distillation
rate and volume of distillate collected were studied. Studies with different mesh size powders and
different quantities of alkali showed that the mesh size is important for maximum recovery with alkali.
since, 0.392 per cent arecoline was obtained for a 20 mm mesh powder , compared to 0.512 per cent
for a 30 mm mesh powder with same quantity of alkali. Studies on the effect of distillation rate
indicated that a rate of 10 ml per min and a distillate volume of 150 ml were optimum for maximum
recovery of arecoline. It was found that there was significant and progressive in the recovery of
arecoline extracted up to six extractions with 30 mm mesh powder sample.

Hu et al. (2009) described the development of nine microsatellite loci from A. catechu. The
number of alleles per locus ranged from 5 to 15. The expected and observed heterozygosities ranged
from 0.71to 0.94 and from 0 to 0.88, respectively. All microsatellite loci, except for AC30,
significantly deviated from Hardy–Weinberg equilibrium possibly due to artificially selected

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.5

cultivation or the existence of excessive null alleles. No linkage disequilibrium was observed from
pairwise comparisons of loci, except for AC06 and AC08.

Senthil amudhan (2010)Reported that ripe arecanut showed high arecoline content than unripe
arecanut. Moderate arecoline content was recorded in at semi ripe stage (6 month) in arecanut. With
respect to arecanut maturity stages, tender nut stage (6-7 month) may be ideal stage for arecanut
chewing purpose which also showed lower arecoline content and this stage may also be ideal for
optimum extraction of polyphenol and CT for pharmacological uses.

Jantaratet al.(2013) used high-performance liquid chromatographic (hplc)with UV detection

method for determination of arecoline and reported that the contents of arecoline in unripe and ripe
areca nuts were 0.1434 ± 0.0016 and 0.0944 ± 0.0002 %w/w of dried seed powder, respectively and
percent recovery was 103.23 ± 5.76 % indicating high precision and accuracy of the method.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 III MATERIALS AND METHODS

This chapter explains the information concerning methodology that was used in conducting the
experiment. It contains a brief description of location of the experimental site, characteristics of soil,
climate, materials, layout and design of the experiment, land preparation, manuring and fertilizing,
transplanting of seedlings, intercultural operations, harvesting, data recording procedure and statistical
analysis etc. which are presented as follows.

3.1. Experimental site

The research work relating to determine the genetic diversity of arecanut was conducted at
Central plantation crops research institute, Regional station, Vittla during 2018.

3.2 Geographical Location of the experimental site

The experiment was carried out at of Central plantation crops research institute, Regional
station, Vittla, which is situated in the Western Ghats and represents the coastal zone (Zone-10) of
Karnataka and lies at 12°766’North latitude and 75° 122’ East longitude with an altitude of 100 m
above mean sea level.Vittla lies in zone-10 of agroclimatic zones of Karnataka. The average rainfall of
area is 2380.72 mm and distributed over a period of six to eight months (April - December) with peaks
during June to September. The meteorological data for the period of experimentation was recorded at
the meteorological observatory of Central plantation crops research institute, Regional station, Vittla.

3.3 Soil characteristics of experimental site

The experiment was conducted in red sandy loam soil. The physical and chemical composition
of soil is presented in Appendix-I.

3.4 Design and layout of the experiment

The experiment was laid out in Randomized Complete Block Design (RCBD) with three
replications. The genotypes were distributed into the every plot of each block of the prepared layout of
the experiment. The fifteen genotypes of the experiment were assigned at random into plots of each
replication. The distance maintained spacing row to row 2.5 m and plant to plant 2 m. The distance
maintained between two blocks was 1 m.

3.5 Planting materials

Fifteen genotypes of arecanut were used for the present research work. The purity and
germination percentage were leveled as around 100 and 80 respectively. The genetically pure and
physically healthy seeds of these genotypes were collected from the Department of Genetics and Plant
Breeding and from local market. The experimental genotypes are presented in Table 1.

3.6 Palm selection for recording observation

Four palms per replication from each germplasm will be recorded.

Table1.Name of fifteen arecanut genotypes used in the present study.

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.7

 Sl. No. Germplasm/Accession No. Germplasm/Accession Name

1 VTL 107 Boragari

2 VTL 108 Jorepakri

3 VTL 109 Kalirhat

4 VTL 111 Bakwabari

5 VTL 112 Rajarhat

6 VTL 113 Goralbari

7 VTL 133 Keri-MO

8 VTL 143 Thirthagundi

9 VTL 152 Vajrahalli

10 VTL 159 Boko

11 VTL 160 Marigaon

12 VTL 161 Makaria

13 VTL 162 Barnihat

14 VTL 163 Amchup

15 VTL 164 East Garuhills

3.6 Harvesting

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 Nuts were picked on the basis of horticultural maturity, size, colour and age being determined
for the purpose of consumption as the nut grew rapidly and soon getbeyond the marketable stage,
frequent picking was done throughout the harvesting period. Nuts were collected with sharp knife and
care was taken to avoid injury of the palm.

3.7 Data recording

Data were recorded on the following parameters from the studied plants throughout their life
cycle. Therecordeddata were on the individual plant basis.

3.7.1. Morphological data

Morphological data were analysed based on the visual observation on a group of plants or parts
of plants. Measurement on a number of individual plants or parts of plants were also used for
morphological data.

3.7.2 Yield and yield contributing data

The data on different yield contributing characters are recorded as follows.

3.7.2.1 Days to first male flowering

The number of days required for first male flowering was counted forthree replications
separately and average data was recorded.

3.7.2.2 Days to first female flowering

The number of days required for first female flowering was recorded for threereplication
separately and average data was recorded.

3.7.2.3 Palm length (m)

Palm length measured in meter in main palm and average data was recorded.

3.7.2.4 Number of nodes per palm

The number of nodes per palm was counted and average data was recorded.

3.7.2.5 Branches per palm

Branches per palm per palm were counted and average data was recorded.

3.7.2.6 Nut length (cm)

Nut length was measured in 3-5 nuts of different plants in cm and average data was recorded
during nut harvest for vegetable use.

3.7.2.7 Nut diameter (cm)

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.9

 Nut diameter was measured in 3-5 nuts of different plants in cm and average data was recorded
during nut harvest for vegetable use.

3.7.2.8 Number of nut per plant

The number of nut per plant was counted and average data was recorded.

3.7.2.9 Weight per nut (g)

Weight of 3-5 nuts of different plants during harvest for vegetable use was measured in gram
(g).

3.7.2.10 Yield per plant (kg)

Weight of edible nutnuts of selected plants from each accession was weighted in kilogram (kg).

3.8 Statistical analysis

Mean data of the characters were subjected to multivariate analysis. Univariate analysis of the
individual character was done for all characters under study using the mean values and was estimated
using MSTAT-C computer programme. Duncan’s Multiple Range Test (DMRT) was performed for all
the characters to test the differences between the means of the genotypes. Mean range and co-efficient
of variation (CV %) were also estimated using MSTAT-C. Multivariate analysis was done by
computer using GENSTAT 5.13 and Microsoft Excel 2000 software through four techniques viz.,
Principal Component Analysis (PCA), principal Coordinate Analysis (PCO), Cluster Analysis (CA)
and Canonical Analysis (CVA).

3.8.1 Estimation of genotypic and phenotypic variances

Genotypic and phenotypic variances were estimated according to the formula of Johnson et al.
(1955).

a. Genotypic variance

Where, MSG = Mean sum of square for genotypes

MSE = Mean sum of square for error, and

r = Number of replication

b. Phenotypic variance

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Where, = Genotypic variance,

= Environmental variance = Mean square of error

3.8.2 Estimation of genotypic and phenotypic co-efficient of variation

Genotypic and phenotypic co-efficient of variation were calculated by the following formula
Burton (1952).

Where, GCV = Genotypic co-efficient of variation

PCV = Phenotypic co-efficient of variation

δg= Genotypic standard deviation

δp= Phenotypic standard deviation

3.8.3 Estimation of heritability

Broad sense heritability was estimated (Lush, 1943) by the following formula, suggested by
Johnson et al. (1955).

3.8.4 Estimation of genetic advance

The expected genetic advance for different characters under selection was estimated using the
formula suggested by Lush (1943) and Johnson et al. (1955).

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.11

3.8.5 Estimation of genetic advance mean’s percentage

Genetic advance as percentage of mean was calculated from the following formula as proposed
by Comstock and Robinson (1952):

3.8.6Estimation of simple correlation co-efficient

Simple correlation co-efficient (r) was estimated with the following formula (Singh and
Chaudhary, 1985; Clark, 1973).

Where,∑ = Summation

x and y are the two variables correlated

N = Number of observation

3.8.7 Path co-efficient analysis

Path co-efficient analysis was done according to the procedure employed by Dewey and Lu
(1959) also quoted in Singh and Chaudhary (1985) using simple correlation values. In path analysis,
correlation co-efficient is partitioned into direct and indirect independent variables on the dependent
variable.

In order to estimate direct & indirect effect of the correlated characters, say x1, x2 and x3 yield
y, a set of simultaneous equations (three equations in this example) is required to be formulated as
shown below:

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Where, r’s denotes simple correlation co-efficient and P’s denote path co-efficient (Unknown). P’s in
the above equations may be conveniently solved by arranging them in matrix from.

Total correlation, say between x1 and y is thus partitioned as follows:

Pyx1 = The direct effect of x1 on y.

Pyx2rx1x2 = The indirect effect of x1 via x2 on y.

Pyx3rx1x3 = The indirect effect of x1 via x3 on y.

After calculating the direct and indirect effect of the characters, residual effect (R) was calculated by
using the formula given below (Singh and Chaudhary, 1985):

Where, P2XY = (R2); and hence residual effect,

Piy = Direct effect of the character on yield

Riy = Correlation of the character with yield

3.8.8 Estimation of Genetic Diversity

The genetic diversity among the genotypes was assessed by Mahalanobis’s(1936) general
distance (D2) statistic and its auxiliary analyses. The parents selection in hybridization programme
based on Mahalanobis’s D2 statistic is more reliable as requisite knowledge of parents in respect of a
mass of characteristics is available prior to crossing. Rao (1952) suggested that the quantification of
genetic diversity through biometrical procedures had made it possible to choose genetically diverse
parents for a hybridization programme. Multivariate analysis viz. Principal Component analysis,
Principal Coordinate analysis, Cluster analysis and Canonical Vector analysis (CVA), which quantify
the differences among several quantitative traits, are efficient method of evaluating genetic diversity.
These are as follows.

3.8.8.1Principal Component Analysis (PCA)

Principal component analysis, one of the multivariate techniques, is used to examine the
interrelationship among several characters and can be done from the sum of squares and product
matrix for the characters. Therefore, principal component were computed from the correlation matrix
and genotype scores obtained from the first components (which has the property of accounting for
maximum variance) and succeeding components with latent roots greater than the unity . Contribution
of the different morphological characters towards divergence is discussed from the latent vectors of the
first two principal components.

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.13

3.8.8.2 Principal Coordinate Analysis (PCO)

Principal coordinate analysis is equivalent to principal component analysis but it is used to

calculate inter-unit distances. Through the use of all dimensions of P it gives the maximum distances
between each pair of the n point using similarity matrix, Digby et al. (1989).

3.8.8.3 Clustering

To divide the genotypes of the study into some number of mutually exclusive groups clustering
were done using non-hierarchical classification. Starting from some initial classification of the
genotypes into required groups, the algorithm repeatedly transfers genotypes from one group to
another so long as such transfers improve the criterion, the algorithm switches to a second stage which
examine the effect of swapping two genotypes of different classes and so on.

3.8.8.4 Average Intra-cluster Distances

The average intra-cluster distances for each cluster was calculated by taking possible D² values
within the member of a cluster obtained from the Principal Coordinate Analysis (PCO). The formula
used was D²/n, where D² is the sum of distances between all possible combinations (n) of the genotype
included in the cluster. The square root of the average D² values represents the distances (D) within
cluster.

3.8.8.5 Canonical Vector analysis (CVA)

Canonical vector analysis (CVA) finds linear combination of original variabilities that
maximize the ratio of between group to within group variation, thereby giving functions of the original
variables that can be used to discriminate between the groups. Thus, n this analysis a scres of
orthogonal transformation sequentially maximizing of the ratio of among groups to the within group
variations. The canonical vector are based upon the roots and vectors of WB, where W is the pooled
within groups covariance matrix and B is the among groups covariance matrix.

3.8.8.6 Selection of varieties for future hybridization programme

Divergence analysis is usually performed to identify the diverse genotypes for hybridization
purposes. The genotypes grouped together are less divergent among themselves than those, which fall
into different clusters. Clusters separated by largest statistical distance (D2) express the maximum
divergence among the genotypes included into these different clusters. Variety (s) or line(s) were
selected for efficient hybridization programme according to Singh and Chuadhury (1985). According
to them the following points should be considered while selecting genotypes for hybridization
programme:

1. Choice of cluster from which genotypes are selected for use as parent (s)

2. Selection of particular genotype(s) from the selected cluster(s)

3. Relative contribution of the characters to the total divergence

4. Other important characters of the genotypes

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 IV RESULTS AND DISCUSSION

The experiment was conducted to perform the diversity analysis of different genotypes of
arecanut (Momordica charantiaL.) using yield contributing and morphological traits. This chapter
comprises the presentation and discussion of the findings obtained from the experiment. The data were
collected from the seedling stage to after harvest stage..The data pertaining to ten morphological
characters and ten yield contributing characters have been presented and statistically analyzed with the
possible interpretations given under the following headings.

4.1 Morphological characterization

Phenotypic expression of morphological characters in arecanut genotypes showed wide range

of variation under studied. The morphological traits of all cucurbitaceous species was reported by
Hutchinson et al. (1954). In the present study, only the characters showing variation are explained.
Phenotypic traits like seed color is the effect of genetic factor exhibited to synthesis or pigment in seed
coat. Above all the described characters are stable and are not influenced by environment. Thus,
selection on the basis of these traits will be effective.

4.1.1Leaf characters

The leaf characters of 15 genotypes were studied based on their number of lobes, depth of
lobbing and length of leaf blade (Table 3). The lobes were characterized based on their number and
depth.

The leaves of seven genotypes (VTL 111, VTL 114, VTL 133, VTL 140, VTL 111, VTL 112
and VTL 1115) had 5 lobes and the leaves of eight genotypes (VTL 112, VTL 131, VTL 132, VTL
139, VTL 141, VTL 1110, VTL 113 and VTL 114) had 7 lobes. Depth of lobing of leaves was
classified as shallow, medium and deep. VTL 112, VTL 141 and VTL 131 showed shallow depth
lobed leaves. VTL 114, VTL 132, VTL 139, VTL 140, 44 VTL 1110, VTL 112, VTL 113 and VTL
1115 genotypes showed medium depth lobed leaves. Deep lobed leaves were found in VTL 111, VTL
133, VTL 111 and VTL 114 genotypes.

Short leaf blade (<6cm) leaves were found in VTL 112, VTL 141 and VTL 1110 (Table 3).
Medium length (>6-9cm)of leaves were found in VTL 114, VTL 131, VTL 139, VTL 140, VTL 111,
VTL 112, VTL 113 and VTL 114 genotypes. The leaves having long (>9cm) type of leaf blades were
found in VTL 111, VTL 132, VTL 133 and in VTL 1115 genotypes. De wilde and Duyfjes (2002)
were found deep and palmate 5-9 lobed leaves in M. CharantiaL.

4.1.2 Nut character

The nut characters in study were classified in nut colour and nut texture. Different nutcolor was
found in two stages. One stage is marketable maturity color where the nuts showed light green color
genotypes (VTL 112, VTL 114, VTL 133 and VTL 141), green color genotypes (VTL 111, VTL 131,
VTL 132, VTL 139, VTL 111, VTL 111 and VTL 114) and dark green color genotypes (VTL 140,
VTL 112 and VTL 113). Another stage was ripening stage where yellow color genotypes (VTL 111,
VTL 114, VTL 132, VTL 133, VTL 139, VTL 112, VTL 113 and VTL 1115) and orange color
genotypes (VTL 112, VTL 131, VTL 141, VTL 111 and VTL 114) were found. Two types of nut
texture were found in the genotypes. Nut ridge were of two types, continuous and discontinuous.
Continuous ridge was found in the genotypes VTL 131, VTL 132, VTL 133, VTL 140, VTL 141, VTL
111 and VTL 114) and discontinuous type of ridge was found in the genotypes VTL 111, VTL

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.15

112,VTL 114, VTL 1110, VTL 113, VTL 114 and VTL 1115). Medium number of tubercles was
found in genotypes VTL 132, VTL 133, VTL 111, VTL 112 and many tubercles found in Genotypes
VTL 111, VTL 114, VTL 131, VTL 139, VTL 140, VTL 141, VTL 1110, VTL 112, VTL 113, VTL
114 and VTL 1115).

4.1.3 Ovary character

Two type ovary of arecanut genotypes were found on the basis of length. One was short (1.5-
2.5 cm) type which was found in the genotypes VTL 132, VTL 139, VTL 140, VTL 141, VTL 113 and
VTL 114.Another was long (>2.5 cm) type genotypes VTL 112, VTL 114, VTL 131, VTL 133, VTL
1110, VTL 111 and VTL 113).

4.1.4 Seed character

Genotypes showed morphological variation in seed character. Seed indentation and seed colour
are the two main characters. Seed indentation was small type, medium type and large type. Small type
indentation was found in VTL 139, VTL 141, VTL 1110, VTL 112, VTL 113 and VTL 114 genotypes.
Medium type indentation was found in VTL 112, VTL 132, and VTL 133and VTL 111 genotypes.
Large type indentation was found in VTL 111, VTL 114, VTL 131 and VTL 140 genotypes.

Three types of seed color were found, light brown, brown and dark brown. The seeds of genotypes
VTL 111, VTL 132, VTL 111 and VTL 114 showed light brown color. Genotypes VTL 114, VTL
133, VTL 140, VTL 141 and VTL 112 showed brown color seed and VTL 112, VTL 131, VTL 139,
VTL 1110, VTL 113 and VTL 1115 showed dark brown color seed.

4.2Genetic variability, heritability and genetic advance

Variability and estimation of genetic parameter in order to different plant characters are
discussed below. The mean performance of different traits of 15 genotypes of arecanut is calculated.

4.2.1 Palm length

For all genetic parameter among fifteen genotypes had adequate variation. There were
significant differences in palm length among the genotypes. The maximum value of palm length (4.32
m) was recorded in VTL 112. This genotype was statistically similar with genotypes VTL 111, VTL
112, VTL 114, VTL 133, VTL 139, VTL 140, VTL 111 and 14. The minimum value(3.28) of palm
length was found in VTL 1115. Robinson and Decker walters (1997) were also found similar
significant variation for palm length. The phonotypic expression of plant characters depend on the
interaction between genotypic characters and environment. The more environmental effect inhibits the
expression of genetic characters. The phenotypic co-efficient of variation was higher than genotypic
one which indicated more influence of environment. Less difference between PCV (10.9%) and GCV
(8.88%) took the advantage of palm length at final harvest. High values of heritability and low values
of genetic advance and genetic advance in percent of mean have recorded for palm length, indicating
that this character was controlled by non-additive gene effects. So, selection of this character wouldn’t
be effective. The result of Chowdhury and Sharma (2002) contradict with the present findings where
the high heritability is coupled with high genetic advance for this character.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

4.2.2 Branch per palm

Number of branch per palm in arecanut genotypes indicated significant variations. The
maximum number of branches was 23.67 and the minimum number of branches was 13.89.The mean
value was 17.33 for number of branches. VTL 141 showed that maximum number of branches pursued
by genotypes VTL 111, VTL 112, VTL 114, VTL 131, VTL 132, VTL 133, VTL 140, VTL 1110,
VTL 111, VTL 112, VTL 113, VTL 114 and VTL 1115. The minimum branches per palm (13.89)
recorded in VTL 139.There was minor environmental influence on the exposure to the character,
because the difference between phenotypic (20.69) and genotypic (20.27) co-efficient of variation was
very close to each other. Suggesting environmental influence is minor on the expression of genes
controlling this trait. So, selection based upon phenotypic expression of this character would be
effective for the improvement of this crop. High heritability with low genetic advance and moderate
genetic advance in percent of mean indicates non-additive gene action controlling this trait. Variation
in branch per palm was also observed in snake gourd genotypes by Banik (2003).

4.2.3 Nodes per palm

Significant differences in nodes per palm were found from 55.00 to 84.33 among the
genotypes. The mean value was recorded 65.42 for nodes per palm. The maximum number of nodes
per palm observed in genotype VTL 111followed by genotypes VTL 141 and VTL 1115. The
genotypic (97.00) and phenotypic (101.07) variance with phenotypic co-efficient of variation (15.37%)
and genotypic co-efficient of variation (15.05%) were indicated the minor environmental impact on the
expression of the character. The heritability estimates for this trait was high (95.98), genetic advance
was (19.88%) and genetic advance in percent of mean (30.38) were found moderate, this trait was
governed by non additive gene. High heritability and low genetic advance for this character was also
observed by Banik (2003).

4.2.4 Days to first flower (male)

The variance due to days to first flowering showed that the genotypes differ significantly. The
range of days to first male flowering were varies from 42.77 days to 51.89 days with mean value 49.77
days (Appendix V). The genotypic co-efficient of variation and phenotypic co-efficient of variation
were 6.61 and 6.72 respectively. The phenotypic variance and genotypic variance suggested minor
influence of environment on the expression of genes controlling the trait. Therefore, selection based
upon phenotypic expression of this character would be effective for the improvement of the crop.A
similar finding was reported by Singh et al.(2002) in ridge gourd.

4.2.5. Days to flower (female)

Significant variation was found for days to female flowering and it ranged from 46.44 DAT in
VTL 111 to 58.33 DAT in VTL 131 with the mean value 53.60 days (Appendix V). The genotypic and
phenotypic variance was 21.96 and 22.06 respectively. The PCV was 8.76% and the GCV was 8.74%..
The phenotypic variance and genotypic variance advised less environmental influence on the
expression of genes governing days to flower (female). Many author also found similar result (Miah et
al. 2000; Sharma et al. 2000).The heritability estimates was high (99.56%) with the low genetic
advance (9.63%) and genetic advance in percent of mean 17.97%, indicating this trait was controlled
by non additive genes. Guffar (2008) support the findings. Plate 6and Plate 7 illustrates the
morphological variation in female and male flower.

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.17

4.2.6. Nut Length

The nut length was noticed as 12.57 cm with a range of 7.67 cm to 17.02cm. The genotype
VTL 111 showed the maximum nut length and the genotype VTL 132 showed the minimum nut
length. The genotypic variance (7.69) and phenotypic variance (7.95) with the co-efficient of variation
genotypic (22.06) and phenotypic (22.43) were closed to each other, indicating minor environmental
influence on this character that would be effective for improvement of this crop. High heritability
estimates 96.76% with low genetic advance (5.62%) over percent of mean (44.71%) (Table 6) indicate
that effective selection may be made for nut length. Sharma et al. (2000), Krisna Prasad and Singh
(1994), Hormuzdi and More (1989) were reported the similar findings for arecanut.

4.2.7 Nut Breadth

The mean nut breadth was 3.83 cm with a range of 2.92 cm (in genotype VTL 1115) to 4.81 cm
(in genotype VTL 139) (Appendix V). Sharma et al. (2000), Krisna Prasad and Singh (1994),
Hormuzdi and More (1989) were found similar results. The genotypic variance (0.51) and phenotypic
variance (0.56) with co-efficient of variation genotypic (18.69%) and phenotypic (19.56%) was found
respectively were closed to each other, indicating minor environmental influence on this character that
would be effective for the improvement of arecanut. High heritability estimates (91.28%) with low
genetic advance (1.41%) over moderate percent of mean (36.78%) indicate that effective selection may
be made for nut breadth. High heritability coupled with low genetic gain for this character was
observed by Sahaet al. (1992).

4.2.8Nut yield (kg) per plant

The nut yield per plant was found 2.27 kg in genotype VTL 111 which is highest and the
lowest was recorded 0.73 kg in genotype VTL 132 with the mean value 1.61 kg (Appendix V). The
genotypic variance (0.38) and phenotypic (0.41) variance (Table 6) suggested less influence of
environment on the expression of the genes controlling this character which gives idea about selection.
Estimation of high heritability (93.28%) with low genetic advance (1.23%) and high genetic advance
of percent (76.10%) revealed that this character was governed by additive gene and provides
opportunity for selecting high valued genotypes for breeding program. Abusaleha and Dutta (1990)
found similar result for this character in arecanut.

4.3 Correlation studies

Determination of correlation co-efficient was provided the information how yield depends on
different yield contributing characters. The correlation co-efficient, values of ‘r’ and the parameter
correlated were shown in table 5.Simple correlation was divided into phenotypic (directly observed)
and enotypic (inherent association between character).

4.3.1 Palm Length

Palm length had non significant and positive correlation with branch per palm (0.399 and
0.372), node per palm (0.215 and 0.194), number of nuts per plant (0.267 and 0.259) at genotypic and
phenotypic level (Table: 5). This finding was supported by Abusaleha and Dutta (1988). Palm length
of arecanut had non significant and negative correlation with days to first male flower (-0.230 and -
0.205), days to first female flower (-0.080) and (-0.073), nut length (-0.209 and -0.83), Nut breadth (-
0.114 and -0.094), nut weight (-0.249 and -0.231) and nut yield per plant (-0.075 and -0.037) at
genotypic and phenotypic level (Table: 4).

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Table 2. Genotypic correlation coefficients among different pairs of yield and yield contributing
characters for different genotypes of arecanut

Traits B/V N/V DFFM DFFF FL FB FW NF/P F/P

VL 0.399 0.215 0.230 -0.080 -0.209 -0.114 -0.249 0.267 -0.075

B/V 0.798** -0.842** -0.491 -0.042 -0.064 -0.092 0.389 0.110

N/V -0.915** -0.676** 0.118 0.080 0.049 0.449 0.244

DFFM 0.830** -0.262 -0.389 -0.082 -0.615* -0.338

DFFF -0.296 -0.545* 0.120 -0.717 -0.174

FL 0.528* 0.581* 0.042 0.679**

FB 0.387 0.545* 0.517*

FW 0.032 0.888**

NF/P 0.323

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.19

4.3.2 Days to first male flowering

Days to first flowering male had significant positive correlation with days to first female
flowering (0.830 and 0.822) at genotypic and phenotypic level (Table:5). This character also
significant negatively correlated with number of nut per plant (-0.615 and -0.603) at genotypic and
phenotypic level. Days to first male flowering also non significant negatively correlated with nut
length (-0.262 and -0.254), nut breadth (-0.389 and -0.367), nutweight(-0.082 and -0.081) and the
significant negative correlation found with number of nut per plant (-0.615 and -0.603) at genotypic
and phenotypic level (Table: 4). Khan et al. (2008) reported almost similar result in case of pointed
gourd.

4.3.3 Days to first female flowering

Days to first female flowering had non significant negatively correlated with nut length (-1.296
and -0.292) and significantly negative correlated with nut breadth (-0.545 and -0.532) and number of
nut per plant (-0.717 and -0.709) at genotypic and phenotypic level. It also negatively correlated with
nut per plant (kg) (-0.174 and -0.172) at genotypic and phenotypic level (table: 4). Guffar (2008) also
found similar result in case of arecanut.

4.3.4 Nut character

Significant positive correlation present in nut yield with nut characters such as nut length
(0.679), nut breadth (0.517) and nut weight (0.888) had a high degree of association (Table: 4). This
showed that with the increase of nut length, nut breadth, nut weight altimately yield per plant will be
increased. Miah et al. (2000); noted that nut yield in arecanut showed significant positive association
with average nut weight, nut breadth and number of nodes per palm in genotypic and phenotypic
correlation with days to male flowering. Sarkeret al. (1999); studied correlation of 16 divergence types
of pointed gourd indicated that nut weight, nut diameter and number of primary branches per plant
were positively and significantly correlated with yield per plant followed by nut weight and nut
diameter had maximum positive direct effects on yield.

4.4 Path analysis

Path analysis showed that the cause and effect situation of dependent and independent variable.
For example yield considered as dependent variable and palm length, days to first flowering and nut
characters are independent variable. Path analysis gives the original pictures of inter relationship
betweenyield and yield attributing characters. Path co-efficient analysis was showed direct and indirect
effects of different characters on yield of arecanut in Table 5.

4.4.1 Palm length

Path co-efficient analysis in table (5) indicated that palm length had direct and positive (0.051)
effect on yield. Palm length had indirect positive effect on yield via branch per palm (0.030), node per
palm (0.016), nut length (0.066), breadth (0.002), Number of nut per plant (0.089). Besides negatively
indirect effect via days to first male flower (-0.013), days to first female flower (-0.089), nut weight (-
0.176) and nut per plant (-0.076) presented in Table (5). Shah and Kale (2002) reported similar result
with the present study and they stated that palm length had positive direct effect on yield per plant.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Table 3. Path coefficient analysis showing direct and indirect effects of different characters on
Nut/Plant (kg) ofarecanut

Nut/Plant (kg)
Branch/Palm
Palm Length

Nut Breadth

Nut Weight
Nut Length
Characters

Node/Palm

correlation
Genotypic
Nut/Plant
No. of
(cm)

(cm)

with
(m)

Palm Length 0.051 0.030 0.016 -0.066 0.002 -0.176 0.089 -0.075
(m)
Branch/Palm 0.020 0.076 0.063 -0.013 0.001 -0.065 0.129 0.110
Node/Palm 0.010 0.061 0.078 0.037 -0.001 0.035 0.149 0.244
Days to 1st -0.012 -0.065 -0.072 -0.083 0.007 -0.058 -0.204 -0.338
flower (male)

Days to 1st -0.004 -0.038 -0.053 -0.093 0.010 0.085 -0.238 -0.174
Flower
(Female)

Nut Length -0.010 -0.003 0.009 0.315 -0.009 0.411 0.014 0.679**
(cm)
Nut Breadth -0.006 -0.005 0.006 0.166 0.018 0.274 0.181 0.517*
(cm)
Nut Weight -0.013 -0.007 0.004 0.183 -0.007 0.708 0.010 0.888**

No.of 0.014 0.030 0.035 0.013 -0.010 0.023 0.332 0.323

Nut/Plant

* and ** significant at 1per cent and 5 percent respectively

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.21

4.4.2 Days to first male flowering

Days to first male flowering had direct positive effect on yield of arecanut (0.056). It also had
indirect negative correlation with yield via palm length (-0.012) branch per palm (-0.065), node per
palm (-0.072) and nut weight. Besides positive indirect correlation with yield via days to first female
flower (0.092), nut length (0.083), nut breadth (0.007) showed in Table (5). Li et al.(1997) were found
negative correlation of days to first male flowering with yield.

4.4.3 Days to first Female flowering

Days to first female flowering had positive direct effect on yield of arecanut (0.111) which
influence to genotypic correlation with nut per plant (-0.174). It also had indirect negative effect with
palm length (-0.004), branch per palm (-0.038), node per palm (-0.053) and number of nut per plant (-
0.238). It had indirect positive effect on yield via days to first male flower (0.047), nut length (0.093),
nut breadth (0.010), nut weight (0.085) Table 5. Rastogi et al. (1990) showed that days to female
flowering had positive and direct effect on yield, which is supported by present findings.

4.4.4 Nut character

Nut length had positive direct effect (0.315) on yield per plant of arecanut genotypes. It had
indirect positive effect via nut weight (0.411), number of nut per plant (0.014) and node per palm
(0.009) (Table 5).Zhang et al. (1999) stated that average nut weight, number of nuts per plant and nut
length had direct effect on yield of arecanut genotypes. There was a significant and positive correlation
between yield and yield attributing characters among indigenous and exotic arecanut cultivars (Rajput
et al., 1991).

4.5 Multivariate analysis

4.5.1 Cluster analysis

The experiment was conducted to investigate the genetic diversity of fifteen genotypes of
arecanut. The genotypes were divided into four cluster according to D2 analysis(table 7).The cluster III
had maximum number of genotypes (7) followed by cluster I which had 4 genotypes. Cluster II and IV
had two genotypes. Remarkably cluster I had (VTL 114, VTL 112, VTL 113 and VTL 1115) where as
cluster II had (VTL 141 and VTL 111). Furthermore cluster III had (VTL 111, VTL 112,VTL
132,VTL 133,VTL 139,VTL 140 and VTL 1115), cluster IV showed two genotypes (VTL 131 and
VTL 1110). Clustering was done at random that indicate a broad genetic base of the genotypes.
Genetic variability in arecanut was also found by Prasad et at. (2001).

4.5.2 Principal component analysis (PCA)

Proper idea about genetic divergence is an important tool for breeding program. The diversity
analysis is useful to determine the magnitude of divergence among population (Murthy and Quadri,
1966). Principal component analysis was studied with fifteen genotypes of arecanut. First three Eigen
values for three principal co-ordination axes of genotypes accounted for 79.68% variation.Based on
principal component scores I and II obtained from the Principal component analysis, a two-
dimensional scatter diagram (Z1-Z2) using component score I as X axis and component score II as Y
axis was Constructed, which has been presented in Figure 1.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Table 4. Distribution of genotypes in different clusters

Cluster Number of genotype Genotypes

I 4 G 3, G 12, G 13, G 14

II 2 G 9, G 11

III 7 G 1, G 2, G 5, G 6, G 7, G 8, G 15

IV 2 G 4, G 10

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.23

Figure 1. Scatter distribution of fifteen genotypes of bitter gourd based on their principal
component scores superior with cluster.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

4.5.3 Principal coordination analysis

Principal coordination analysis (PCO) indicated that the maximum inter genotypic distance
between genotype VTL 131 and VTL 132 (1.637), the minimum distance was showed between
genotype VTL 111 and genotype VTL 140.There was moderate level of variation present among
fifteen genotypes of arecanut due to low inter genotypic distance. The maximum intra-cluster distance
was presented in cluster I (0.938) which had four genotypes (VTL 114, VTL 112, VTL 113, VTL
114). The minimum intra-cluster distance was recorded in cluster II which containing two genotypes
(VTL 141,VTL 111). Khan et al. (2008) reported twelve clusters in pointed gourd. Quaruzzamanet al.
(2008) reported six clusters in ridge gourd.

4.5.4 Non-hierarchical clustering

Fifteen (Momordica charantiaL.) genotypes were divided into different group for non-
hierarchical clustering. These finding ensures the clustering pattern of the genotypes gained by PCA. It
indicated that cluster 1 contained four genotypes, cluster II and IV contained two genotypes and cluster
III presented seven genotypes of arecanut. From cluster mean, cluster II had the maximum mean value
for seven characters namely palm length (4.21 cm), Branch per palm (23.29) node per palm (83.28),
nut length (13.57), nut breadth (4.21 cm), nut weight (58.04 gm), Number of nut per plant (83.61) and
nut yield per plant (1.96 kg). This cluster mean gives idea about the cluster II could be used for future
hybridization program for palm length (m), branch per palm, node per palm, nut length (cm), number
of nut per plant and nut per plant (kg). Cluster III had required for days to first flower male (51.05) and
days to first flower female (54.87). Cluster IV had highest mean value for nut weight (101.57) and
days to first flower required (56.39) days, cluster III and cluster IV had moderate mean value for all
character. These genotypes of cluster could be used for future hybridization program. Singh et al.
(2013) reported that contribution of the characters to the divergence in arecanut.

4.5.5 Conical variate analysis

Conical variate analysis (CVA) was done to calculate the inter-cluster distance. The data
presented intra and inter-cluster distance (D2) values. In this study the inter-cluster distances were
more than the intra-cluster distances. It proved that the wide range of genetic variability among
genotypes of arecanut. On the basis of intra and inter cluster (D2) value, the close cluster of cluster 1
was cluster III (9.790) and distant cluster was cluster II (25.16). figure 2 Cluster II consists of nearest
cluster with) cluster was cluster III (30.55). With the help of D2 values within and between clusters, an
arbitrary cluster diagra was constructed, which showed the relationship between different genotypes.
Diagram also showed the intra and inter cluster distance of fifteen genotype of arecanut. Shanmugam
and Rangasamy(1982) stated that genotypes distributed in different clusters are sign of broad genetic
base of diversity.

4.5.6 Selection of genotypes as parent for hybridization programme

Genetically dissimilar parent selection is the fundamental works for hybridization programme.
So the genotypes were chosen according to specific trait, maximum heterosis could be shown in
offspring from the crosses between genetically diverse parents. On the basis of cluster mean and
agronomic performance the genotype VTL 111 for maximum palm length, branch per palm, early
flowering, nut length, number of nut and nut weight. VTL 113 for maximum nut length, nut weight
and nut per plant (kg) were found promising. Therefore considering group distance and other
agronomic performance the inter genotypic crosses between VTL 141, VTL 111, VTL 131 and VTL
113 and also other improved variety and might be suggested for future hybridization program.

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.25

Table 5. Cluster mean values of 10 different characters of 15 genotypes of arecanut

Parameters I II III IV
Palm Length (m) 3.87 4.21 4.02 3.79
Branch/Palm 15.77 23.39 16.67 16.71
Node/Palm 61.80 83.28 62.62 64.61
Days to 1st flower 50.52 43.28 51.05 50.33
(male)
Days to 1st Flower 53.78 46.00 54.87 56.39
(Female)

Nut Length (cm) 13.57 13.57 11.46 13.49

Nut Breadth (cm) 4.12 4.21 3.50 4.00
Nut Weight 65.17 58.04 42.97 101.57
No. of Nut/Plant 27.75 33.61 26.46 27.77
Nut/Plant (kg) 1.82 1.96 1.18 2.30

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

Figure 2. Diagram showing intra and inter cluster distance of fifteen genotype of bitter gourd

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.27

 V SUMMARY AND CONCLUSION

The experiment was conducted at college of horticulture, Mudigere with fifteen genotypes of
arecanut during October 2015 to February 2016. Seeds were sown on pots. The seedling transplanted
into the main field in Randomized complete Block Design (RCBD) with three replication. As most of
the morphological traits are not influenced by the environment, transfer of those characters is fully
governed by genes and could be use in hybridization program. From this point of view, morphological
characterization of leaf, nut, flower and seed characters were performed. In order to investigate yield
potential, genetic variability, character association, genetic divergence of fifteen genotypes data were
recorded on yield contributing parameter such as palm length, branch per palm, node per palm, days to
first flower (male), days to first flower (female), nut length, nut breadth, nut weight, number of nut per
plant, nut per plant (kg) under study.

The genotypic and phenotypic variance along with co-efficient of variation for both (genotypic
and phenotypic), showed adequate variation present among genotypes for all characters. In case of
palm length, nut weight, number of nut per plant, nut yield per plant indicated more environmental
influence for expression of the trait. On the other hand branch per palm, node per palm, nut length, nut
breadth indicate minimum genotypic and phenotypic variance difference suggesting additive gene
action for the expression of the characters.

Correlation co-efficient among the characters were used to investigate the relationship between
yield and yield attributing traits. Most of the character showed higher genotypic correlation co-
efficient than phenotypic correlation co-efficient which gives idea about the relationship between the
characters under study. The significant positive correlation was found between yield and nut length
and nut weight at genotypic and phenotypic level. There was non-significant positive correlation
between yield and branch per palm, node per palm, number of nut per plant at genotypic and
phenotypic level. On the other hand non-significant negative correlation was found between yield and
palm length, days to first flower male and female at genotypic and phenotypic level.

Considering the magnitude of cluster mean and yield attributing traits, it can be concluded that
the performance of the VTL 141 for palm length, branch per palm, early flowering and no. of nut per
plant was excellent. VTL 131 for highest nut length and nut weight, VTL 111 for early flowering and
nut yield per plant, VTL 113 for maximum number of nut per plant and higher yield were found
promising. Therefore VTL 131, VTL 141, VTL 111 and VTL 113 considered as improved genotypes
for high yielding genotypes and might be suggested for future hybridization program.

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 VI REFERENCES

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Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes

 APPENDICES

A. Physical composition of the soil.

Soil separates %

Sand 36.90

Silt 26.40

Clay 36.66

Texture class Clay loam

Kiran Kuma.D.G., M.Sc.(Hort.), 2018.31

B.Chemical composition of the soils.

Sl. No. Soil characteristics Analytical data

1 Organic carbon (%) 0.82

2 Total N (kg/ha) 1790.00

3 Total S (ppm) 225.00

4 Total P (ppm) 840.00

5 Available N (kg/ha) 54.00

6 Available P (kg/ha) 69.00

7 Exchangeable K (kg/ha) 89.50

8 Available S (ppm) 16.00

9 pH (1:2.5 soil to water) 5.55

10 CEC 11.23

Morphological and molecular characterization of arecanut (Areca catechu L.) genotypes