The genus Garcinia L. belongs to the family Clusiaceae, having approximately
two hundred species distributed in Asia. Thirty six species are found in India
among them G. gummigutta and G. indica are more important ones.
G. indica is an evergreen tree with drooping branches found endemic in
tropical rain forests of the western ghats restricted to Konkan region of Maharashtra.
Some industries have started extracting Hydroxy-Citric Acid (HCA) from the rind
of the fruit which is an important constituent used as an hypocholesterolaemic
and anti-obesity agent (Heymsfield et al., 1998).
Fruit extract is traditionally used as soft drink with attractive colour, recently
has shown to have antioxidant activity (Mishra et al.,
2006). The fruit rinds of G. indica contain cyanindin-3-glucoside
and cyanindin-3- sambubioside (Nayak et al., 2010).
Various pigments of G. indica were screened for inhibition of enzymes
hyaluronidase and elastase, thus, contributing to antiageing activity (Sahasrabudhe
and Deodhar, 2010).
Since 1995, in K.E.Ts V.G. Vaze college, a study was intiated to identify
the early fruiting, high yielding and HCA rich clone of G. indica. Method
of clonal propogation for elite species was standardize for G. indica
(Chabukswar and Deodhar, 2006). G. indica is
dioecious in nature and lot of morphological variations in leaf shape, branching
pattern, thickness of fruit rinds, colour of fruit, tree height was observed
in various locations of G. indica. Hence systematic study was undertaken
to collect the plants from various populations, and to see whether the polymorphism
also exists at genetic level. Various populations were screened with molecular
markers (Thatte et al., 2012) and karyomorphological
studies at different locations were also undertaken. The present study aims
at karytomorphological study at different locations.
There are only two reports on karyomorphology of G. indica. Much earlier
in 1949, while study the karyomorphology of some fruit trees, Krishnaswamy and
Raman reported the chromosome number of G. indica to be 2n = 54. Razdan
(1972) reported diploid chromosome number of G. indica to be 2n =
MATERIALS AND METHODS
Collection of plant material: Seeds of G. indica were collected from various regions (Fig. 1), Deorukh (Dist. Rantagiri), Otawane (Dist. Sindhudurg) and Diveagar (Dist. Raigad) in June 2009, 2010, 2011 and 2012. The seeds were germinated in plastic trays in polyhouse of K.E.Ts V.G. Vaze college, Mulund (E.), Mumbai- 400081, Maharashtra.
Chromosome preparation: The mitotic index was determined by analyzing
maximum number of dividing cells in the root tip of the seedling of G. indica.
|| Geographic locations of G. indica populations used
in this study.
Indicates collection sites
The maximum number of dividing cells were observed in the morning at 10:45
am; so root tips were excised at this time and pretreated with p-Dichlorobenzene
for 3 h at 4°C. Then roots were transferred in distilled water and kept
at 4°C for 1 h. The root tips were fixed in 10% acetic acid for ten minutes,
washed and hydrolysed in 1N HCl at 60°C for 5 min in thermostat. Squash
were made in 2% propionic orcein. Chromosomes images were taken with Olympus
microscope reflected light fluorescence attachment (Olympus CXRFA -2 model)
equipped with Pinnacle software.
Chromosome analysis: Measurement of chromosomes were done by Biovis image plus V4.11 software. The length of chromosomes were measured in micrometer. The number of chromosome, ploidy level, karyotype formula, ratio of longest to shortest chromosome, mean and median of long arm length, mean and median of short arm length, total chromosome length, total form percent, mean centromeric index and symmetry classes determined from seven well spread metaphase cells. Chromosome homology was assigned according to similarities in length, morphology and centromere position. Chromosome nomenclature based on the centromere location followed that proposed by Levan, i.e., Metacentric (M), Submetacentric (SM), Telocentric (T) and Acrocentric (AC). Chromosomes were paired with their respective chromosomes and were arranged as 1 to 54 in descending order of length. When all measurements had been compiled, the total chromosome length, total form percent (TF%) and Centromeric Index (CI) were calculated. Karyotype formula of the population investigated were summarized.
In present communication, the plant material was collected from different Konkan
region of Maharashtra. At least seven well spread metaphase cells were considered
for karyomorphological studies. The karyotypes of all studied plants correspond
to the different formula. The study was based on three population of G. indica
maintaining chromosome number 2n = 54 (Fig. 2a-c).
The karyotype formula as well as respective karyotypes (Fig. 3a-c)
obtained and the parameters analysed were summarised in Table
For constructing the karyotype and calculations, the chromosomes were arranged
in order of decreasing size (Fig. 4a-c).
Chromosome number (2n), karyotype formula, the Longest Chromosome Length (LCL),
the Shortest Chromosome Length (SCL), long arm/short arm ratios (L/S) and centromeric
index (CI) of G. indica are listed in Table 1.
All the populations of G. indica were diploid, with 2n = 54.
The somatic chromosome in all the population was constant 2n = 54 but differ in some morphological aspects. In Deorukh population (Fig. 2a) chromosome number was 54. Karyotype analysis showed one pair of satellite, one pail of secondary constriction and one pair of asymmetric chromosome. Otawane population (Fig. 2b) from Sawantwadi region was showing similar ploidy level having one pair of satellite chromosome but differ in pattern of secondary constriction. The secondary constriction appeared in rod shape. It also showed one pair of asymmetric chromosome. Where as Dive agar population ( Fig. 2c) from Raigad district was showing 54 chromosomes with three pairs of satellite chromosomes, one pair with secondary constriction and one pair with asymmetric chromosome.
The observed value of total chromosome length of complement in Deorukh population
(120.1384 μm) and Otawane population (118.3909 μm) were found comparatively
lesser than that of population from Diveagar (131.0864 μm).
|| The karyotype formula and the parameters analysed
||Microphotographs of well spread mitotic metaphase plates of
G. indica (a) Accession from Otawane, (b) Accession from Deorukh
and (c) Accession from Diveagar
Shortest chromosome were observed in Deorukh (1.24998 μm) and in Otawane
(1.1443 μm) which was found comparatively lesser than that of Dive agar
(1.42757 μm). Longest chromosome found in Dive agar (4.0934 μm) was
comparatively higher than Deorukh (3.4027 μm) and Otawane (3.4748 μm)
The karyotypes investigated can be summarized as follows:
||The composition of the karyotype gives ground to distinguish
||Deorukh population 2n = 54 = 14M+ 34SM+ 2SMSAT+2SMSC+2
||Otawane population 2n = 54 = 14M+ 34SM+ 2SMSAT+2SMSC+2
||Diveagar population 2n = 54 = 14M+ 30SM+ 4SMSAT+2SMSC+2TSAT+2
||Heteromorphic chromosomes pairs were established in all the
||Karyotypes of G. indica (a) Population of Deorukh (b)
Population of Otawane and (c) Population of diveagar
|| Ideogram of G. indica (a) Deorukh population, (b)
Otawane population and (c) Diveagar population
From family Clusiaceae in genus Clusia, n = 30 seems to be constant
for the genus (Cruz et al., 1990). Robson
and Adams (1968) suggested the basic chromosome number for Hypericum
to be x = 8 or 9. For other allied species in family Clusiaceae, Callophyllum
and Mesua it is reported to be n = 8 and 16, respectively and the same
count applies to Genus Garcinia.
Razdan (1972) determined the chromosome number of various
species of genus Garcinia. He has reported haploid number for Garcinia
is n = 24. Species like G. morella and G. cowa are diploids representing
2n = 48. for G. in G. cambogia, G. spicata and G. parviflora are
triploids that is 2n = 72 whereas G. mangostana is 2n = 96. The exceptions
are G. livingstonii 2n = 56 and G. xanthochymus 2n = 80. It is
possible that these might be hybrids.
In present study all the populations of G. indica were diploid, 2n = 54.
There are several reports of chromosome polymorphism observed among populations
in several species which reported both numerical and structural variations.
Chengqi (2008) studied somatic chromosome number of
Allium przewalskianum in different populations. They screened 302 individuals
from 43 locations, 90 were diploid 2n = 16 and 212 were tetraploid 2n = 32.
Li (2005) studied chromosome number of Aster ageratoides
an endemic to china and related if geographically. Most of the populations were
hexaploid (2n = 6x = 54) an occupied an extensive area of south west china to
eastern chile. And diploid and tetraploids were less frequent and limited to
There are several reports including a species in different populations e.g.,
Melilotus officinalis (Pavlova and Toshewa, 2004),
Cicer arietinum (Kordi et al., 2006)
and Impatiens balsamina (Momtaz et al.,
2007) exhibited morphological variations in chromosomes. Pavlova
and Toshewa (2004) studied karyomorphology of Melilotus officinalis in
populations of Bulgaria. All M. officinalis populations were diploids
(2n = 16). On the basis of karyomorphology, the populations can be divided into
3 groups. The group one comprised of 10 metacentric, 4 submetacentric chromosomes
with a pair of satellite was on submetacentric chromosome. In group II also
there were 10 metacentric, 4 submetacentric chromosomes but the satellite was
attached to the metacentric type chromosomes. The group III was having 8 metacentric,
4 submetacentric chromosomes. In this group also the satellite was associated
with metacentric chromosomes and there was one pair of intercentric chromosome
(Pavlova and Toshewa, 2004). They also observed deviation
in macromorphology of these plants in group III. The plants were shorter with
less number of inflorescence, larger legumes and hair on lower leaflet surface.
They called this taxon as variety arenaria it was locally endemic suggested
that it should be under control and protected.
In present study also out of three populations, two populations Deorukh and
Otawane had 14 metacentric, 34 submetacentric chromosomes, pair of asymmetric
chromosome and one unique pair of submetacentric satellite chromosome. But in
Otawane population the secondary constriction appeared to be rod shaped (Fig.
2b). Diveagar populations showed 14 metacentric, 30 submetacentric chromosomes.
Satellite appeared on 3 pairs of chromosomes. Out of them two satellite appeared
on submetacentric chromosomes (Fig. 4c) In addition to this
there was a pair of telocentric chromosomes with satellites which was unique
to this populations. They also have comparatively longer chromosomes (Table
Razdan (1972) reported karyomorphology of three different
cytotypes of G. indica. In cytotype one there were four pairs of satellite
chromosome while in cytotype two had three satellite and one chromosome pair
with secondary constriction which matches with our observations. But a pair
of heteromorphic chromosome was not reported earlier.
G. indica being dioecious insect or cross pollinated plant was expected
to show lot of morphological variation in canopy shape, shape of leaves, branching
pattern, thickness of fruits etc. hence the plants from various locations were
studied morphologically. Also percentage of polymorphism was studied using RAPD
and ISSR markers (Thatte et al., 2012). To our
surprise most of the populations like Dapoli, Chiplun, Shirode of Sawantwadi
showed only 4-5% polymorphism. In comparison the plants in Diveagar populations
were significantly taller having significantly longer leaf lamina. And these
plants also showed higher degree polymorphism (8-10%), showing some unique bands
characteristics to this populations (Thatte et al.,
2012). These plants also differ karyomorphologically.
The plants of G. indica located in small pocket of Diveagar can be considered as separate taxon. As the plant is highly endemic, there is urgent need for conserve the cytotypes.