Genetic diversity of spine gourd cultivars

SPINE GOURD Momordica dioica
Vernacular name: Kantola, Kakrol
Submitted by
S.ADHIYAMAAN (2017603401)
I-M.Sc.,VEGETABLE SCIENCE
DEPT. OF VEGETABLE CROPS
HC & RI, TNAU, CBE.- 641 003

Origin
Spine gourd is believed to be originated
in the tropics of the old world.
Centres in India:
West Bengal and Karnataka are two
major Indian states that grow Kantola
commercially.
Kantola is largely cultivated in Malda and
Nadia districts of West Bengal.

MEDICINAL VALUE
• Possess hypoglycemic, hepatoprotective, gastroprotective and
ulcer healing activities, analgesic, expectorant, nematocidal,
antiallergic, antimalarial, antifeedant, antibacterial and antifungal
activity

Distinct Character
• Anthesis at evening 6-7 PM
• Starts flowering 30–40 days after planting depending upon prevailing
weather conditions and lasts up to 4–5 months.
• Individual fruit weight is 5- 20 gm
• Density of seeds in fruits
• It is tolerant to pumpkin caterpillar, gall fly and root knot nematode

Spine gourd genotype RMF 37 identified as variety “Indira Kakoda 1” by Central
Variety Release committee (CVRC) for 5 states i.e. Chhattisgarh, Uttar Pradesh,
Jharkhand, Orissa and Maharastra during Group Meet held at PAU, Ludhiana
(Punjab) from May 5-6, 2006.
It is resistant to all major pests, green fruit are attractive dark, green and contain 12-14
% protein. Its fruits are dark green coloured.
It yields 0.8-1.0 q/ha during 1st year, 1.0-1.5 q/ha in 2nd yearand 1.5–2.0 t/ha in
third year after planting.
Varieties: Indira Kankad 1

• Developed through clonal selection at CHES, Bhubaneswar.
• High yield (4–5 kg/ plant) and high market preference.
• Its vines are thin and spreading which grows well on 3-line wire-trellis
system.
• Moderately resistant to anthracnose and downy mildew in fields, and is
moderately susceptible to angular leaf blight and pumpkin caterpillar.
Varieties: Arka Neelachal Sree

Climate: Spine gourd is a warm season crop and can be successfully cultivated
in the plains and hills of sub-tropical and tropical regions. Plenty of sunshine and
low humidity are ideal conditions for its growth. Warm humid climate, 25-40◦C
Soil: A well-drained sandy loam soil with a pH 5.5 - 6.5. Rainfall- 150-250 cm
Sowing Time: Kantola can be grown both as a summer crop and a rainy season
crop. Sowing time for kantola crop in tropical plains is January –February for
summer crop and July-August for rainy season crop. Sowing time in tropical
hills is April.
Seed Rate: Approximately, 1-2 Kg seeds are required for sowing one acre of area.

• Propagation: Seed, Tuber and Cuttings
• Seed: dormancy (5-6 months), dip in H2O -24hrs
• 30 ◦C better germn(Ali et al, 1991)
• Plants from seeds can not be determined (1:1 male &female)
• By Tuber: dormancy(4-5 months), healthy plants, 2-3 yrs old plants &120-150g, 2 buds /piece
• Treated with Thiourea 87.5% sprouting than control(18.3%) Panda et al ,(1993).
• Planting : Sep-Oct or Feb-Mar
• Ram et al.(2002) Feb-Mar irrigated cond. &June-July rainy season
PROPAGATION

Adventitious root tubers from vine cuttings
Efficient method for multiplication
3000- 5000 tubers /ha
Spacing = (2 X 1)m

• By Cuttings: Dark green cuttings from 2-3 years old form basal or terminal portion
with 2-3 nodes planted in sand bed after rooting planted in main field
• Tripathy et al, 1993: Terminal cutting is the best
• Sahu et al, 1995 no significant effects on growth and devolp. of due to type of
cuttings and no of nodes in the cuttings
• Ahmad et al ,1992 obtained 93% rooting, 17 roots/cutting comprising one axillary bud
and a mature diploid leaf when treated with 1500pppm of IBA in 5s and planted in
mixture of soil: sand: compost (1:1:1)
• Grafting: C.moschata, C.trifolia, F1 hybrids of C.moschata x C.maxima
PROPAGATION

Field Preparation: Field is well prepared by 2 – 3 ploughing followed by tilling and
levelling of the soil. Well-rotten organic manure or FYM (farm yard manure)@10-15
tonnes/acre is incorporated with the top soil during the last ploughing to enhance soil
fertility.
Spacing: Recommended spacing to be followed (if staking is not practiced) is, 1-2 meter
between two ridges and 60-90 cm between two plants. If staking is practiced, this spacing
may be reduced according to your cultivation requirements.
Sowing Depth: Seeds are sown by placing two or three seeds deep into the pits prepared
on raised beds at 1-2 cm depth. If more seeds/pit is used, thinning should be done after
the sprouting of seedlings.

• Application of ethephon to male plants of kakrol did not affect the plants at
any level of concentration tested while application of AgNO3 (400 ppm)
produced the highest number of bisexual flowers per vine. (Ali et al. 1991)
• Foliar sprays AgNO3 (400 ppm) at preflowering stage could induce 70–90
% hermaphrodite flowers in M. dioica. (Rajput et al. 1994)
Use of PGRs for sex alteration

• A high parthenocarpic fruit set (70%) was observed in an inter-specific
hybrid between M. dioica and M. cochinchinensis when the F1 was
pollinated with pollen from M. cochinchinensis. - Bharathi et al. (2012)
• Induction of parthenocarpy in M. dioica with pollen of related taxa
(M. charantia and Lagenaria leucantha) and mixture of the pollens from
these two species. - Singh (1978)
• The parthenocarpic fruit setting was higher with the stimulus of
extraneous pollen (66% against 36%), compared to natural pollination
Parthenocarpy

Harvesting: Kantola is harvested when their
fruits are still young and tender.
 Harvesting at every alternate day or at 2-3
days intervals is recommended to avoid
losses due to over-sized and over-mature
fruits
Yield: An average yield of 650 g/plant is obtained
under good cultural practices. That is, approximately
5 t/ac.

M. subangulata subsp. renigera
• Wild vegetable native to Assam-
Myanmar region.
• Distinct dark dots at the base of pale
yellow to nearly white corolla.
• The fruits of M. subangulata are
narrowed distinctly towards tip.

• In M. subangulata there are two subspecies, subangulata with
longitudinal ridges, no spines, surface totally smooth,
• And subsp. renigera with tubercles present and in longitudinal rows,
surface more or less spinescent if ridges are present;

Momordica sahyadrica
• Native to Assam- Myanmar region
• Tendrillar, dioecious, perennial climbers.
• Flowers are bright yellow with a narrow greenish yellow base.
• Fruits broadly ellipsoid, or ovoid to fusiform, or with round base and rostrate
apex.
• Turning bright orange on ripening, densely clothed with soft short spines 2-4
mm long; pulp sweet when ripe, carmine red.

Momordica muricata
• Annual climber with unbranched glabrous tendrils, monoecious.
• Stem pubescent to glabrescent.
• Flowers yellow.
• Male flowers solitary, (1.5) cm long peduncle, bearing near the apex a
sessile.
• Female flowers on 5-15 mm long.

Fruits-of-Momordica-spp- a) M. balsamina; b) M. charantia;
c)M. cochinchinensis d) M.dioica e) M. sahyadrica; f) M. subangalata

REFERENCES
• http://www.agrisk.umn.edu/
• https://www.echocommunity.org/en/

Research articles
• 1.Genetic relatedness (diversity) and cultivar identification.pptx
• 2.In vitro rooting studies in spine gourd.pptx
• 3.Variability and correlation studies in spine gourd.pptx

Genetic relatedness (diversity) and cultivar identification by randomly
amplified polymorphic DNA (RAPD) markers in teasle gourd
(Momordica dioica Roxb.)
M.G. Rasul a,*, M. Hiramatsu b, H. Okubo b
a Department of Genetics and Plant Breeding, Bangabandhu Sheikh Mujibur Rahman
Agricultural University, Salna, Gazipur 1706, Bangladesh
b Laboratory of Horticultural Science, Faculty of Agriculture, Kyushu University 46-01,
Fukuoka 812-8581, Japan
Received 29 May 2005; received in revised form 28 May 2006; accepted 23 October 2006

INTRODUCTION
• The tetraploid and triploid forms in M. dioica are also reported in
same population collected from Khashi and Jayantia Hills of Assam,
India (Roy et al., 1966; Agarwal and Roy, 1976).
• Moreover, unique markers developed by different techniques may
serve a very essential tool for certification of plant materials as true to
type, for paternity tests and in future, for genetic mapping.

INTRODUCTION
• RAPD marker (Williams et al., 1990) has proven quite useful in
genetic study of many plant species (Baral and Bosland, 2002; Teng et
al., 2002; Levi and Rowland, 1997).
• RAPD is a dominant marker, does not require target sequence
information for design of amplification of primers.

OBJECTIVES
• (a) To establish a DNA extraction technique,
• (b) To obtain DNA fingerprints,
• (c) To estimate genetic similarity and diversity among 30 teasle gourd
accessions collected from different regions of Bangladesh.

MATERIALS AND METHODS-Plant materials
The tuberous root of 30 accessions
(29 accessions of M. dioica Roxb.
and 1 accession of Momordica
cochinchinesis Spreng.) were
collected only from different
representative agro-ecological region
of Bangladesh in 2001
Accession numbers of teasle gourd and collection sites in Bangladesh

DNA extraction and quantification-CTAB method with
some modifications.
1. Aprox. 70 mg of fresh leaf was ground in a mortar with 50 ml of liquid
nitrogen.
2. Add 1.5 ml of extraction buffer 1, to the ground powder, again grinded .
The samples were centrifuged for 5 min at 12,000 rpm and the
supernatant was discarded. (This step was done twice)
3. Then the sample was re-extracted with 300 ml extraction buffer 2 and
200 ml extraction buffer 3

some modifications.
4. The sample was mixed vigorously with vortex machine and incubated in a hot
water bath at 60.8°C for 10 min.
5. A solution of chloroform and iso-amyl alcohol (24:1, 500 ml) was added to the
suspension and mixed gently and the samples were kept for 15 min at room
temperature.
6. After that the sample was centrifuged for 5 min at 12,000 rpm at 20.8°C. The
aqueous phase (500 ml) was transferred to a new eppendorf tube and 350 ml
ice-cold isopropanol was added and mixed gently and it was precipitated at
_20.8°C for at least 30 min (over-night is also ok).
Contd..

some modifications.
7. The sample was then centrifuged at 12,000 rpm for 5 min at 4°C. Then
800 ml of 70% ethanol was added to the pelleted DNA and again
centrifuged at 12,000 rpm for 5 min at 4°C.
8. The pellet in eppendorf tube was dried in a vacuum dessiccator for 7 min
and resuspended with 50 ml TE (Tris– HCl, EDTA).
9. One microlitre of RNase (0.5 mg/ml) was added to DNA suspension and
incubated in a water bath at 60°C for an hour.
Contd..

some modifications.
10. The sample was then mixed with vortex for a short time and again
incubated in a water bath for 20 min at 60.8°C.
11. The DNA concentration was determined in comparison with l- DNA
digested with HindIII (lHindIII ladder) by electrophoresis in 1.5%
agarose gel in 1_ TBE (Tris–borate–EDTA) buffer.
12. The DNA solution was stored at _20.8°C until analyzed.
Contd..

MATERIALS AND METHODS-Oligonucleotide primers
• RAPD common primers
were obtained from DNA
oligomer (12 mer) set A
(Wako, Tokyo). The
sequences of the primers,
GC contents and Tm (°C)
ratio
Primer name, sequence, total bands, percentage and polymorphic RAPD bands for each primer among 30 teasle gourd accessions

PCR–RAPD assay
• As usual, Each reaction mixture was laid out with one drop of mineral oil
to prevent evaporation.
• It programmed for 1 cycle of 30 s at 94°C for pre-denaturation followed
by 45 cycles of 94°C for 30 s, 42°C for 2 min and 72°C for 3 min for
denaturing, annealing and DNA extension, respectively. The last cycle
was followed by incubation at 72°C for 7 min followed by holding
indefinitely at 4°C.

PCR–RAPD assay
• The PCR products were loaded in 1.5% agarose gel and separated by
electrophoresis with 1 % TBE (Tris–borate–EDTA) buffer at 100 V for 40 min.
• The PCR product loaded in the last lane of the previous gel and first lane in the
subsequent gel were duplicated for providing an internal control in scoring
RAPD bands from more than one gel.
• The gels were stained with ethidium bromide (0.5 mg/ml) viewed under
ultraviolet transilluminator and photographed with Polaroid camera.
• Molecular sizes of the amplification products were estimated using l-DNA
digested with HindIII (lHindIII ladder).
Contd..

RAPD band scoring and genetic analysis
• RAPD bands were scored as present (1) or absent (0) in each genotype for each
set of primers
• The band scoring data were used to calculate the similarity (s) and distance (d)
matrices with the following equations: similarity, s = 2Nxy/(Nx + Ny) and
genetic distance, d = -1 x log(s), where
• Nxy = number shared bands between the genotypes ‘x’ and ‘y’,
• Nx = total number of bands found in genotype ‘x’, and
• Ny = total number of bands found in genotype ‘y’
• (Nei and Li, 1979).

RAPD band scoring and genetic analysis
• The similarity value lies between 0 and 1.
• A similarity value 1 indicates complete or 100% genetic similarity between
two accessions while a similarity value 0 indicates no genetic similarity.
• A dendrogram was constructed on the basis of distance matrix unweighed
pair group method with arithmetic average cluster analysis, UPGMA
method using the software PHYLIP Version 3.57c (Felsenstein, 1995).

RESULT - RAPD analysis (band polymorphism)

The highest number of bands was produced by
the primer CMNA 62

Accession identification (fingerprinting)
Accessions identified with two or more primersPrimers with specific unique bands (presence or absence of
single band) with their molecular size

Genetic relationship and diversity among the accessions
• Similarity matrix among 30 teasle gourd accessions generated using Nei and Li index

Cluster analysis
• Dendrogram of 30 teasle
gourd (Momordica spp.)
accessions constructed using
a UPGMA cluster analysis
method based on Nei and Li
(1979) genetic distance
value by PHYLIP program.
There was no relation between geographical origin and genetic diversity.

DISCUSSION
• The high polymorphism in our study was due to using a high number
of primers and proved the effectiveness of RAPD techniques in
Momordica.
• Thus the duplication of names could be discriminated clearly and
independently by one or two primers as also recommended

DISCUSSION
• Founder effects meant that only a small fraction of genetic variation of
a parent population or species present in the small number of founder
member of a new colony or population.
• Acc 30 (showed much genetic distance) could be used as one parent in
enriching genetic variability (large fruit size, submergence tolerance)
of teasle gourd through interspecific hybridization

CONCLUSION
• Genetic fingerprinting by RAPD technique has proven to be a useful
method for identification and classification of teasle gourd accessions and
cultivars
• Since this is the first report of using RAPD markers in teasle gourd, the
genetic similarity value developed in this study provides teasle gourd
breeders with a starting point in designing crosses to increase the genetic
diversity of their material.

In vitro rooting studies in spine
gourd (Momordica dioica Roxb)
Dipali V. Ghive, R. B. Ghorade, R. P. Khedekar,G. S. Jeughale and N.W.Raut
Dr. Panjabrao Deshmukh Krishi Vidyapeeth, Akola (M.S.) India
Asian J. of Bio Sci. (2006) Vol. 1 No. 2 : 146-148

INTRODUCTION
• Propagation by seed and cuttings is not common in Kartoli.
• The highly cross pollinated dioicious nature of Kartoli exhibit genetic variation
for morphological and growth parameters.
• Tubers are suitable for propagation but it have some limitations, first is dormancy
and sprout only at the onset of monsoon secondly multiplication rate of tuber is
very low.
• High multiplication ratio can be achieved by micropropagation technique, which
enables rapid multiplication of disease pest free elite plants within short space and
time.

MATERIALS AND METHODS
• The explants of spine gourd i.e., shoot tip
and axillary buds were washed under tap
water and surface sterilized after washing in
0.1% Teepol detergent for 2 minutes and
followed by 0.1% HgCl2 treatment for 2
minutes.
• A single shoot from well established culture
were transferred to Rooting Media
RM1 (MS+IBA 0.2 mg/l)
RM4 (MS +IBA 2.0 mg/l)
RM6 (MS+AdSO4 80 mg/l + IBA 1.0 mg/l)
RM8 (MS+AdSO4 80 mg/l +IBA 3.0 mg/l)

RESUITS AND DISCUSSION
Effect of different concentrations of cytokinins and auxins on per cent rooting

Effect of different concentrations of cytokinins and auxins on number of primary roots

Effect of different concentrations of cytokinins and auxins on root length (cm)

Variability and correlation studies in spine gourd
(Momordica dioica Roxb.)
P BASUMATARY1*, G C BORA2, U C KALITA3, L SAIKIA4 and N C DEKA5
1Regional Agricultural Research Station, Assam Agricultural university, Diphu 782460, India.
2Department of Horticulture, College of Agriculture, A.A.U, Jorhat 785013, India.
3Department of Plant breeding and genetics, College of Agriculture, A.A.U, Jorhat 785013, India.
4Department of Horticulture, College of Agriculture, A.A.U, Jorhat 785013, India.
5Department of Agronomy, College of Agriculture, A.A.U, Jorhat 785013, India.

INTRODUCTION
Correlation studies between yield and other traits of the crop will be
of interest to the breeders in planning the hybridization programme
and evaluating the individual plants in segregating populations.
The characters having high genetic coefficient of variation indicate
high potential for effective selection.

MATERIALS AND METHODS
15 spine gourd germplasm collected from farmer’s field of different parts
The tubers of female and male genotypes (SPG-05 ) were planted at the ratio of 10:1 to ensure better fertilization.
Four cultivars already studied and maintained by All India Coordinated Research Project (AICRP) on
Vegetable crops (VC), Jorhat centre were also used as check varieties in the study.
The experiment was laid out as per Augmented design where check genotypes were replicated thrice
and the test genotypes were kept unreplicated and distributed unequally in the blocks.

RESULTS AND DISCUSSION
Analysis of variance for Plant Characters.
Analysis of variance for Yield and Fruit Characters.

RESULTS AND DISCUSSION
Analysis of variance for Seed and Tuber Characters.
Analysis of variance for Crop Duration and Biochemical Characters.

Genotypic variance, phenotypic variance, genotypic coefficient of variation,
phenotypic coefficient of variation, range and mean of yield and yield
attributing characters of 15 spine gourd germplasm.
high genetic coefficient of variation indicate high potential for effective selection.

Correlation between yield and its attributing characters
Vine length is highly correlated with internode length, fruit diameter and 100 seed weight. Again number of primary
branches was highly correlated with internode length and fruit yield. Internode length highly correlated with fruits per
plant, fruit diameter and fruit yield. Days to 50 percent flowering showed highly positive correlation with total soluble
sugars (%).

CONCLUSION
• This study concluded that there was sufficient genetic variability among the
germplasm lines for most of the characters under study. Hence selection will be
operative for improvement of those characters.
• The significance of positive correlation for yield with number of primary
branches, internode length, fruits per plant, fruit diameter and single fruit
weight indicates that simple selection for any of these characters will result in
considerable improvement of yield in spine gourd.

Genetic diversity of spine gourd cultivars

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