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Karnataka J. Agric. Sci.

, 21(4) (498-502) : 2008

Cytological Analysis of Interspecific Hybrid between Sesamum indicum L X S.
Orientale L. Var . malabaricum

M. S. ANNAPURNA KISHORE KUMAR AND S. C. HIREMATH

Department of Botany
Karnatak University, Dharwad-580 003, India.
Email-annapurnakishore@gmail.com

(Received : November, 2008)

Abstract: The cytology and genome relations studied interspeific hybrid between Sesamum indicum L. and S.orientale Var.
malabaricum Nar. reveal n=13 or 2n=26 chromosome and are diploid on x =13 base number. Morphologically S.orientale
Var.malabaricum is similar to crop species S.indicum cultivated sesame and these two taxa have identical chromosome number
of 2n=2x=26.In the present work S.indicum and S.oriental Var.malabaricum, were intercrossed to establish the genome
relations within the genetic assemblage .The rationale of genome analysis is that the chromosome that pair are closely related
and those that do not pair are not related. The chaismata pairing is a good measure of segment-by-segment homology and
seemingly of nucleotide sequence homology along the length of the intimately paired chromosomes. In a cross of S.indicum
as female and var. malabaricum as male parent the crossability was 40% and large number of F1 hybrids were raised. In the
hybrid mean chromosome pairing of 0.0I+12.6II+0.0III+0.2IV per cell was found. Regular 13 bivalents formation was noticed
in 80% of PMCs. The F1 hybrids were partially fertile and none of F2 seeds germinated. Thus genomes of S.indicum and Var.
malabaricum are similar and homologus. Var. malabaricum forms a member of “AA” genome of S.indicum speices complex.

Key words : Sesamum indicum L., Sesamum orientale, “AA” genome, genome analysis

Introduction wild species possess genes for resistance to viral/fungal disease
and insect pests. Genome analysis is the inference of relationship
Genus Sesamum L. belongs to the family Pedaliaceae among chromosome sets on the basis of the degree of their
and its species are widely distributed in tropical Africa, synapsis. It is possible only when two taxa can be hybridized
Madagascar, Indian subcontinent and some island of Malayan and fertile hybrids obtained. The nature of chromosome pairing
Archipelago.Sesame or Sesamum indicum L. is an important oil can be analysed in hybrids between species. Knowledge of
seed crop, commonly known as til, gingelly, simsim, benniseed, genome relationships between plant species is very useful in
gergelin etc. Sesame is an ancient oil seed first recorded as a planning effective breeding strategies designed to transfer
crop in Babylon and Assyria over 4000 years ago. Sesamum, desirable genes or gene clusters from one species into another,
along with coconut is regarded as the oldest oil-yielding plant there by producing fruitful genomic reconstructions. The genus
known to man. At present in India about 26 million hectares of sesamum require definitive investigations in the direction of
land is under oilseed cultivation. Sesame oil carries a premium genome relationship and phylogenetic studies. In the present
relative to other cooking oils. Sesame oil is esteemed as a food investigation interspecific breeding experiments were conducted
because of its high quality and stability. It possesses resistance to establish the genome relations and to analyze the extent of
to oxidative rancidity. It is also used for anointing, making soup, genome divergence of the genome.
antioxidants, insecticides and medicines. The cytogenetics of
the genus Sesamum is poorly understood and requires more Material and Methods
definitive investigations. Sesamum exhibits two basic Hybridization experiments were conducted in open field
chromosome numbers, x=8 and x=13, based on these two base with potted plants, flower buds destined to open next morning
numbers diploid, tetraploid and octaploid species have been were hand emasculated in the previous evening and bagged.
recognized. Sesame is extremely susceptible to salinity Pollen from the desired male parent was collected and dusted
conditions and water logging. There are a number of serious on the stigmatic surface of the female parent between 7 a.m. to 8
pests and diseases. Aphids, leaf rollers (Antigastra a.m. Pollinated flowers were re-bagged and a label was tied which
catalaunalis), green vegetable bugs and leaf-hoppers are the contained information like date of pollination, time, name of
major pests, Diacrisia obliqua, a hairy caterpillar is capsule parental species etc. Capsule formation and its growth indicate
borer. Fungal diseases are leaf spots caused by Cercospora the success of the cross and the same was inspected after 5 to
sesami Zimm., Alternaria spp., Fusarium oxysporum. 6 days after pollination. In each cross 15-20 emasculated
Schlecht.Ex.Fr., sesami cast. stem and root rot caused by unpollinated flowers were bagged as control. After 20-30 days
Phytophthora and Rhizoctonia spp. Bacterial leaf spot disease of crossing hybrid seeds were collected and were germinated
due to Pseudomonas sesami causes extensive yield loss. Several after 2 to 3 months. All the crossing data like parents, number of
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Karnataka J. Agric. Sci., 21(4) : 2008

flowers pollinated, number of fruit set and genetic marker among plant taxa. Most importantly genome analysis provides
characters of parents and F1 were carefully recorded in the useful information on chromosome paring relationships and
observation book. Pollen fertility experiment was conducted by hence on transferring desirable results from one species to
using Brewbacker and Kwack (1963) medium. another. Genome analysis in broad sense refers to genome
relationships among diploid species and its relationship to
From each plant about 10 pistil were randomly selected polyploid species. Association between chromosomes of the
before pollination and number of ovules/ pistil were counted. parental species resulted in the information of conspicuously
The mean number of well developed seeds/capsule were heteromorphic bivalents suggesting an ancestral relationship
determined. The data was pooled to get mean numbers of ovules between the two species. (Jauhar & Joppa 1996). Identity of wild
/pistil and well developed seeds /capsule from randomly selected progenitor of Sesamum indicum L is highly speculative (Bedigian
plants. Seed fertility of parents and their hybrids were determined 1988), Joshi (1961), Nayar and Mehra (1970), Kobayashi (1993)
by estimating the mean number of ovules / pistil and well and Hiremath S.C. and Patil C.G. (1999,2002) has reviewed the
developed seeds/capsule. present status of genome relations among Sesamum genus.
Meiotic studies were made from young flower buds Hiremath and Patil (1999) obtained successful hybrid
fixed in acetic alcohol (1:3) for 24 hrs. Anthers were dissected from a cross S.indicum X S.mulayanum .Mean chromosome
out from young flower buds using a dissecting binocular pairing of 0.21 I + 12.22 II + 0 III + 0.33 IV per cell was found.
microscope. They were smeared in 1% aceto-carmine on the About 71% of the PMCs showed 13 bivalents. Hiremath and
slide. The slides were made permanent using Celariar's (1956)
Patil (1999) suggested that genomes of these 2 species are
acetic acid butanol series and mounted in euparal. The PMCs
similar and homologous.Genomic symbol AA is proposed for S.
were analyzed from dry permanent slides and all the observations
indicum and S. mulayanum. Further genomes of these two taxa
were carefully recorded. Photomicrographs were taken with 35
have diverged and differentiated through 2 translocations as
mm camera using 8X eye piece and 100X (1.32 N.A.)
evident from 2 quadrivalent formation in the hybrid.F1 hybrids
apochromatic objective. The germplasm of S.indicum were
were partially fertile.
obtained from Dr. Herta Kolberg and Dr. C. Mannheimer, NPGRC,
Windhoek, Namibia and S.orientale . Var. malabaricum were Inthe cross S. indicum X S.orientale,Var. malabaricum
obtained from Dr. Sethupathi, Ramalingam, TNAU, Coimbatore, Sesamum indicum Linn. is used as pistil donor. It is an erect,
India, annual herb, 100-150 cm high, branched at the base, stem
rectangular, pubescent, glandular, leaves heteromorphic,
Results and Discussion
opposite or alternate, petiolate, 3-10 cm long, flowers solitary in
Genus Sesamum has maximum development and the leaf axil , shortly pedicellate, white, pink or purple, some
differentiation in Africa and encompasses about 30 species but
most of them are synonyms of 20 odd species (Ihlenfeldt and
Seidensticker 1968, 1979, Merxmuller 1968, Bruce 1953, Stapf
1906). Currently following species have been recognized :
S.indicum, S.angolense, S.latifolium, S.alatum, S.angustifolium,
S.calycinum (with four sub species) S. parviflorum,
S. abbreviaturm, S.capense, S.Marlothii, S.pedaloides,
S.rigidum, (with two sub species), S.schinzianum S.triphyllum,
S.radiatum, S.Hendelotii, S.mombazense from Africa (stapt 1906,
Bruce 1953, Ihlenfeldt and Seidensticker 1968, 1979, Merxmuller
1968, Seidensticker 1988, Nayar and Mehra 1970). Kobayashi
(1993) has reported S. auriculatam and S.brasiliense from Crete
and Brazil. S.mulayanam , S.laciniatum and S.prostratum are
known to occur in India (Hooker 1885, Nair 1963).

The information about genome relationship in sesame
at intraspecific and interspecific level is meager. The genus
sesamum require definitive investigations in the direction of
genome relationship and phylogenetic studies. Knowledge of
genome relationships between various plant species is of great Figs.1-6. Morphology of parent and F1 hybrid:1.Habit of S.indicum
importance to cytogeneticists, plant breeders, biosystematicists, ;2.Habit of F1 hybrid ; 3.Habit of S.orientale var.malabaricum;
molecular biologists, taxonomists and evolutionists. Genome 4.Flowering twig of S.indicum; 5.Flowering twig of F1 hybrid; 6.
analysis helps in deciphering the cytogenetic architecture of Flowering twig of S.orientale var.malabaricum.
polyploid species and in elucidating phylogenetic relationships
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Cytological Analysis of Interspecific . . . . . . .. .....

erect, much branched annual herb growing to a height of 2 to 5
feet. Stem is quadrangular with a longitudinal groove running
the whole length of each side, tinged with purple, gland dotted
all over and hairy, hairs gland tipped. Leaves heteromorphic,
long petiolate, ovate acute or deeply 3 lobed, margin distinctly
or indistinctly dentate, dark green pubescent. The leaves at top
sub opposite or alternate and reduced in size. Flowers purple,
conspicuous, solitary, short pedicel, with two yellow nectary
glands opens on maturity. Corolla 25-30 cm long light to dark
purple, dark purple crescent present on anterior side and purple
dotted all over. Stamens 4, epipetalous, didynamous, dorsifixed,
a dark purple longitudinal line along the line of dehiscence,
capsule erect, 2.0-2.5 cm long, pubescent with short beak, seeds
black, ovate 0.2cm to 0.3cm long, 1 mm thick distinctly reticulate
rugose and bounded by a raised sharp margin, connecting border
of seed is abrupt (Figs. 3,6,9 & 12).

Figs.7-12. Morphology of parents and F1 hybrid:7.Flower of
S.indicum;8.Flower of F 1 hybrid ; 9.Flower of S.orientale
var.malabaricum;10.Seeds of S.indicum;11.Seeds of F1 hybrid;
12.Seeds of S.orientale var.malabaricum.

times spotted, pedicels usually short with nectarial glands at
the base. Corolla foxglove like, obliquely companulate, limb
sublabiate, lowest lobe is the largest. Filament slender, anthers
dorsifixed, cells parallel, connective gland tipped. Ovary bilocular,
capsule oblong, grooved, beaked, at the apex. Seeds numerous,
obovate, compressed, white in colour, wingless and testa smooth
(Figs.1,4,7 &10).

In the present work the S.orientale Var.malabaricum.
is used as pollen donor. S.orientaleVar.malabaricum, Nar is
wild variety of cultivated sesame. This variety is always found Figs.13-21.Meiosis in parents and F1 hybrid:13.Metaphase–I ,13
only on uncultivated laterite hill tops in a wild state, occurs in bivalents inS.indicum;14.Metaphase–I ,13 bivalents S.orientale Var.
the region of Malabar to Bombay and Central Provinces. This malabaricum;Fig 15-21. Meiosis in F1 hybrid;15.Metaphase –I ,13
wild variety of sesame possesses a unique character to with bivalents;16.Metaphase–I,11II +1IV17.Metaphase –I 9II+2IV
;18.Anaphase –I normal ;19 Anaphase –I with laggards ;20. Anaphase–
stand heavy rainfall. It has been found gregariously in wild II normal; 21.Tetrad normal
state and described as wild variety. This wild variety is an

Table 1 : Morphological characters of parents and their F1 hybrids
Characters S.indicum F1 hybrid S.orientale,Var. malabaricum
Stem Glabrous Glabrous Pubescent
Stem colour Dark green Yellowish Yellowish
Inter nodal length Long Medium Short
Leaf Glabrous Pubescent Pubescent
Leaf glands Absent Present Present
Corolla colour White Pink Pink
Lower lip of flower White Violet Violet
Colour of anther marking White Purple Purple
Style length Long Medium Short
Extra floral nectary Absent Present Present
Seed coat White Black reticulate Black reticulate

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Karnataka J. Agric. Sci., 21(4) : 2008

According to the findings and detail studies of John quantitative characters. Comparative morphological characters
et.al (1950) S.orientale,Var.malabaricum not considered as a of parents and their hybrid are given in table -1 and Figs. 1-6.
valid species but a variety of sesame correctly refered as
S.orientale Var. malabaricum is a weed in malabar region of Comparative meiotic behaviour of parents and hybrid
Kerala. Bedigian and Harlan (1986) considered it is a progenitor : Both the parental species are diploid with 2n=2x=26
of sesame. Nair (1995) belive it to be weedy escape from chromosomes. In female parent S.indicum, the meiotic process
was normal. All the cells analyzed showed regular 13 bivalent
cultivation. Present author (Annapurna L.K, 2003) obtained the
formation (Fig.13). Subsequent meiotic process was normal. In
seed samples of Var. malabaricum from Tamil Nadu Agricultural
the male parent Var.malabaricum, the meiosis was regular.
University, Coimbatore (T.N.) and compared to morphology with
Thirteen bivalent are observed in all the cells screened. Further
S.indicum and S.mulayanum, Var.malabaricum was crossed to
meiotic divisions were normal. (Fig.14 ). In the F1 hybrid mean
S.indicum and S.mulayanum to establish the genomic affinities
chromosome pairing of 0I+12.6II+0.0III+0.22IV per cell was
and resolve the identity of wild progenitor of sesame in her
found. Different chromosome configurations and their
Ph.D work and opined that S.orientaleVar. malabaricum
frequencies are given in table-2. Nearly 80% of the PMCs showed
described by John et al. (1950) is not a variety of S.indicum
regular 13 bivalent in each PMC (Fig. 15) univalents and trivalents
rather it is S.mulayanum itself. Hiremath and Patil (1999) based
were conspicuously absent. One or 2 quadrivalents were also
on morphology and chromosome pairing date of S.indicum X observed in about 17.33% of cells analyzed (Figs. 16 & 17)
S.mulayanum hybrid suggested that S.mulayanum may be the Subsequent meiotic process was slightly irregular. Telophase-I
wild progenitor of cultivated sesame. S.mulayanum is a wild normal and Telophase-II exhibited a micronucleus. Ten percent
species distributed in India and grow gregariously in wasteland of tetrads showed varying number of micronuclei. Pollen fertility
and abandoned field (Nair 1963). in S.indicum was 94% , where as, seed set was 92%, in
In the cross S. indicum X S.orientale,Var. malabaricum var.malabaricum showed 92% pollen fertility, while seed set
about 45 flowers of S.indicum were pollinated with the pollen was about 80%. In the F1 hybrid, 45% of the pollen was assessed
of S.malabaricum. Eighteen capsules were collected with 40% good with 35% seed set. Large number of seeds germinated but
fruit set. In the reciprocal cross 59 flowers of Var. malabaricum none of these survived. The F1 hybrids were partially fertile and
were pollinated with the pollen of S.indicum and 17 hybrid none of F2 seeds germinated. Thus genomes of S.indicum and
capsules were obtained with the fruit set of 32.69%. In S.indicum Var. malabaricum are similar and homologus. Var. malabaricum
X S.orientale Var.malabaricum cross, about 425 seeds were forms a member of "AA" genome of S.indicum speices complex.
obtained from 18 capsules. Out of these only 20 seeds germinated Reciprocal cross was not successful. Hiremath and Patil (1999)
and 12 of them reached to maturity. Seventeen capsules of were not successful in obtaining viable hybrid of of
reciprocal cross Var. malabaricum X S.indicum yielded 560 S.mulayanum X S.indicum .The present author in her Ph.D
seeds and 25 seeds germinated but none of them reached to research work (Annapurna L.K,2003) produced a hybrid between
maturity. Majority of F1 seeds were shriveled and were nonviable. S.mulayanum X S.indicum and studied , crossability was 42%,
Morphological genetic marker characters were used to confirm F1 hybrid mean chromosome pairing of 0.04I+12.5II+0III+0.23IV
the hybrids. per cell was formed. About 82% of the PMCs showed 13
bivalents. This pairing behaviour indicates that genomes of
Comparative morphological studies of hybrid with S.indicum and S.mulayanum are similar and wild S.mulayanum
parents : The F1 hybrids were similar to female parent in glabrous may be the progenitor of S.indicum. The Ph.D work of Annapurna
nature of stem F1 hybrids resembled male parent with regard to L.K (Annapurna L.K, 2003) fully supports the findings of
yellowish stem colour, pubescent leaves, presence of leaf glands, Hiremath and Patil (1999). Cultivated sesame and its progenitor
pink coloured corolla, violet, coloured lower lip, purple marking S.mulayanum grow in the same areas but there is no information
on anther, presence of extra floral nectaries and black reticulate is available on introgression between these taxa. There is an
seeds. They were intermediate with respect to several urgent need to study the ecology and natural distribution of

Table2 : Chromosome pairing in parents and their F1 hybrids

Species/Hybrid Chromosome association No. of cells analyzed Percentage of cells analyzed
I II III IV
S.indicum (Coll No.225) - 13 - - 90 100
X chromosome pairing /cell - 13 - - 90 100
S.orientale,Var. malabaricum - 13 - - 74 100
X chromosome pairing/ cell - 13 - - 74 100
F1 hybrid - 13 - 90 82.57
- 11 - 1 14 12.84
- 9 - 2 05 4.59
X chromosome pairing / cell - 12.6 - 0.22 109 100

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Cytological Analysis of Interspecific . . . . . . .. .....

S.mulayanum. Ease of crossability, formation of 13 bivalents S.indicum, wild species S.mulayanum and S.orientale Var.
and fertility of hybrids in S. indicum X S.orientale malabaricum are similar and homologous. These three taxa
Var.malabaricum , S.mulayanum X S.indicum (Hiremath and belong to "AA" genomic group and form a tight genetic
Patil, 1999) and S.indicum X S.mulayanum (Annapurna, 2003) assemblage within the genus Sesamum. Geographic distributions
cross indicate that S.indicum, S.mulayanum and S.orientale
and morphology of S.orientale Var. malabaricum is identical to
Var. malabaricum are closely related and share a common "AA"
the wild species S.mulayanum. study based on morphology,
genome. Within the diploid species of Sesamum these three
taxa forms a genetic assemblage with similar "AA" genome and crossability, chromosome pairing data in the hybrid and fertility
comparative morphology of S.mulayanum and S.orientale Var. of F1 and F2 reveal that S.mulayanum is the wild progenitor of
malabaricum reveal that they are similar with reticulately rugose oil seed crop S.indicum. Sesame was domesticated through
seed surface in contract to smooth read surface in all the known selection and further cultivation of a large grained smooth
varieties of sesame S.indicum. Genomes of cultivated sesame surfaced genotype of S.mulayanum (Annapurna, 2003).

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