Advances in hybrid seed production of tomato

ADVANCES IN HYBRID SEEDADVANCES IN HYBRID SEED
PRODUCTION TECHNOLOGY OFPRODUCTION TECHNOLOGY OF
TOMATOTOMATO
Speaker:
Akshay Chittora
Ph.D. Horticulture

CONTENTSCONTENTS
• Introduction
• Hand emasculation and pollination
• Use of male sterility
• Biotechnological tools in hybrid seed production
• Major constraints in utilization of F1 hybrids
• Conclusion

TOMATOTOMATO
Common Name :Tomato
Botanical Name :Solanum lycopersicum
formerly Lycopersicon esculentum
Introduced in India :19th
century by Britishers
Chromosome No. :2n = 24
Family :Solanaceae (nightshade)
Centre of origin :Peru (South America)

AREA, PRODUCTION AND PRODUCTIVITY OF TOMATO
AREA
(000' ha)
PRODUCTION
(000’Mt)
PRODUCTIVITY
(Mt/ha)
World 4815.71 163029.746 33.9
China 1000 50000 50
India 882.03 18735.9 21.2
NHB (2014)
Tomato shares 9.4% of total vegetable area and 11.5% of total vegetable
production in India

Historical BackgroundHistorical Background
♦ Among the vegetable crops, first commercial F1 hybrid
of brinjal was released during 1924 in Japan
♦ In tomato first F1 hybrid was developed in 1940 in
Japan
♦ At national level first hybrid vigor was reported in chilli
during 1933 in by IARI, New Delhi
♦ First public sector hybrid developed was Pusa
Meghdoot in bottle gourd in 1971
♦ The first hybrid vegetable seeds (Karnataka
of tomato and Bharat of bell pepper) were

a) Single cross hybrid (AXB)
b) Three way cross hybrid (AXB) X C
c) Double cross hybrid (AXB) X (CXD)
d) Modified single cross (AXA’) X B
e) Double modified single cross (AXA’) X (BXB’)
f) Modified three-way hybrid (AXB) X (CXC’)
g) Top cross hybrid (Inbred X Variety or family etc.)
Types of HybridsTypes of Hybrids

Advantages of Hybrids
 Increased yield
 Early maturity
 Heat & cold tolerance
 Disease and pest resistance
 Better uniformity
 Better fruit quality
 Better transportability
 Better keeping quality

STEPS OF HYBRID SEEDS PRODUCTION
Production of inbred lines
In Self-Pollinated : Pure line
In Cross-Pollinated : Inbred (by selfing)
Testing of combining ability
GCA for additive Gene Actions
SCA for dominant Gene Action
Predictive Information from SCA by
(Single Cross, Double Cross, Three way Cross, Top Cross Poly Cross
and Diallele)
Improvement of inbred lines / varieties
For disease and quality trait
Production of hybrid seed
Types of Hybrids and their Seed Production
Hybridization

The commonly utilized mechanisms for developing
commercial hybrids in vegetables
Mechanism Commercially exploited crops
Hand emasculation + MP Tomato, Eggplant, Okra
Pinching of staminate flowers + MP/NP Cucurbits
Male sterility + MP Tomato, Hot pepper, Sweet pepper
Male sterility + NP Onion, Cabbage, Cauliflower, Carrot,
Radish, Muskmelon, Chilli
Self incompatibility + NP Cauliflower, Cabbage
Gynoecism + NP Cucumber, Muskmelon
PGR & pinching of staminate flowers + NP Summer squash, Winter squash etc.
Hazra and Som (2009)
MP: Manual pollination
NP: Natural pollination

FLOWERING BEHAVIOUR
ANTHESIS DEHISCENCE RECEPTIVITY
OF STIGMA
Starts at 6 AM and
maximum flower
opening till late
morning.
8 AM – 11 AM. 16 hrs before and 5
days after anthesis.

Hand emasculation andHand emasculation and
pollination as hybridizationpollination as hybridization
processprocess
 Plant male and female plants in a separate plots.
 Plant male seedlings 2-3 weeks earlier to obtain adequate
amount of pollen
 Ratio of female to male is generally 4:1
 Spacing
Female plant : 75-90 X 60-75 cm
Male plant : 60 X 45-60 cm
 Roguing at before flowering, early flowering and fruit setting
stage and fruiting stage

Treatments No. of flowers
crossed per
plant
No. of
crossed
fruit per
plant
Fruit set
(%)
Crossed fruit
yield per plant
(g)
Seed yield /
plant (g)
Stage of bud (S)
S1 76.52 20.73 27.12 1549.71 6.19
S2 76.84 40.65 52.92 2558.29 7.75
SEm± 2.15 0.61 1.21 61.08 0.17
CD @ 5% NS 1.83 3.67 183.87 NS
No. of pollinations (P)
P1 77.31 29.26 38.07 1899.80 6.50
P2 76.10 29.69 39.00 1989.04 6.84
P3 76.62 33.11 43.00 2533.16 7.23
SEm± 1.98 0.50 1.05 49.21 0.15
CD@ 5% NS 1.52 3.15 146.15 0.44
(Jolli et al., 2009)
Effect of stage of bud and no. of pollination on different parameters of
tomato hybrid (DTH-1)
S1: <50% bud open, S2: >50% bud open
P1: One time pollination (morning), P2: Two time pollination (morning + evening),
P3: Three time pollination (morning, evening and next day morning)

USE OF MALE STERILE LINES
Types of male sterility found in tomato and there inheritance pattern(Kaul, 1988:
TYPES OF
STERILITY
DESCRIPTION INHERITANCE
Pollen Pollen abortive Monogenic recessive (ms series)
Positional Sigma exerted Monogenic recessive (ps)
Functional Anthers do not dehisce Monogenic recessive (ps 2)
Staminal Stamens absent Monogenic recessive (sl)

sl (Stamenless mutant) in tomato

Natural population
Artificially induced through
• Mutagenesis
•Distant hybridization
• Protoplast fusion
• Genetic engineering

PROCEDURE FOR DEVELOPMENT HYBRID USING
GENEIC MALE STERILE LINES
Parents msms X MsMs
(male sterile) (male fertile)
Msms
(male fertile)
F2 1 MsMs : 2Msms : 1 msms Ratio 3:1
(male fertile) (male sterile)
F1
x

Study done at IARI using male sterile &
male fertile lines in Tomato
• Time saved by male sterile line was 63%.
• Average fruit set by using male sterile line was 79.35%.
• Average fruit set using male fertile line was 65.40%.
• Hybrid seed cost by using male sterile line per kg was Rs. 466/-
• Hybrid seed cost by using male ferile line per kg was Rs. 3691/-
Tewari (1997)

Dhaliwal and Cheema (2008)
Time required for crossing 50 flower buds on male fertile (Ms33 IPA)
and genetic male sterile (ms33 IPA) plants in tomato.

Maintenance of maleMaintenance of male
sterilitysterility• Genic or pollen male sterility
msms (male sterile) x Msms (male fertile)
1 Msms : 1 msms
(rogued out)

Drawbacks in male sterilityDrawbacks in male sterility
• Maintenance (Identification and roguing) is difficult in
pollen abortive (ms) sterility
• ps 2 sterility has variable expressivity and there is
necessity of staminal emasculation
• Outcrossing leading to poor seed set
(0.3-1.5 kg/acre seed as a result of natural outcrossing)
(Yardanov, 1983)

Linkage of ms gene with the marker gene in tomato
Marker geneMarker gene ReferencesReferences
• Potato leaf shape & green
stem colour
•Parthenocarpic fruit
•Enzyme markers
• Purple coloured hypocotyle
•Anthocyanin absent
•Kaul,1988
•Soressi & Salamini,1975
•Tanksley et al.,1984
•Georgiev, 1991
The ms-1035 gene is linked with a recessive marker gene aa
(absence of anthocyanin). Hence, ms-1035 sterile plant can be
identified at seedling stage and fertile plant can be rouged out in
the nursery itself. Moreover, no effect of genes ms1035aa on plant
and fruit characteristics was established.
(Atanassova and Georgiev,

For utilizing functional sterility anther emasculation could
be made easier if ps 2 was combined with short style.
Such flowers can be emasculated without using forceps
(Georgiev and Atanassova 1981)

USE OF CHEMICAL HYBRIDIZING AGENTS (CHA)
Applied chemicals Remark(s) Reference
GA3 Treated plants produced separate
stamens and split pistils
Chandra Sekhar
and Sawhney,1990
Gibberlin synthesis
CCC inhibitors
Selectively inhibited the development
of stamen or msuppressed pollen
Rastogi and Singh,
1988
ABA (Abscisic acid) Selectively inhibited the development
of stamen or suppressed pollen
Rastogi and Singh,
1988
FW-450 (Mendok) Showed promise for commercial
utilization
Moore, 1959
Dalapon Male sterility was induced Brauer, 1959
TIBA (Triiodobenzoic
Acid)
Some degree of male sterility was
inducing
Rehm, 1952
NAA Induced male sterility McRae, 1985

Biotechnological tools useful inBiotechnological tools useful in
Hybrid seed productionHybrid seed production
Tissue culture
 Clonal multiplication via in vitro fertilisation
 Haploid culture
 Protoplast fusion
Transgenic approaches
 for male sterility
 for developing resistant line
 terminator seed technology
Molecular Markers

Application of haploidApplication of haploid
cultureculture
CROP FINDINGS REFERENC
E
Tomato Haploidy has been
successfully used for
developing male sterile
pure line
Zamit et al.,
1980
Schereva et.al.,
1990
42% MS plant+34%
normal plant obtained
by culturing cv.Roma &
MS line with ms 1035
Oankh et al.,
1986

Application of Protoplast fusion
 Through protoplast fusion of Solanum
esculentum with Solanum acuale & S.
tuberosum, cytoplasmic male sterile cybrid
plant with different flower morphology have
been isolated (Melchers et al., 1992)
 Male sterile cytoplasm has been transferred
into S. pennelli & then CMS pennelli has
been successfully crossed with esculentum
(Petrova et al., 1999)

Use of molecUlar markers
 Marker assisted transfer of specific genes/ QTLs
controlling heterosis for desirable traits
 Assessment of genetic diversity
 Establishment of heterotic pools
 Prediction of hybrid performance
 Testing of genetic purity of parental lines and
hybrid seeds

Analysis of Genetic Diversity in 11 Tomato
Varieties using RAPD Markers
Tabassum et al. (2013)
 Twenty arbitrary oligonucleotide primers used in the RAPD‐PCR produced a
total of 584 different marker bands with an average of 29.2 bands per
primer.
 Based on the banding pattern 94.168% polymorphism observed among the
tomato varieties
 The values of pair‐wise genetic distances ranged from 0.1838 - 0.9049,
indicating the presence of wide genetic diversity.

Determination of Genetic Purity of Hybrid Seed in
Watermelon and Tomato Using RAPD markers
Hashizume et al. (1993)

Mechanism Remark(s) Reference
Nuclear male
sterility
Monogenic recessive mutant was utilized to
develop cost effective experimental crosses.
Sawhney, 1997;
Kumar et al., 2001
Auxotrophy A very attractive and feasible model was
proposed utilizing monogenic recessive
nutritional mutants e.g. thiamin dependent.
Barabas, 1991
Incongruity Models to transfer barrier (incompatibility
between pollen and stigma) genes and
corresponding penetration genes were
demonstrated.
Hogenboom et al.,
1978
Commercially unexploited mechanisms for the
development of hybrids in tomato

HYBRID INSTITUTE
Arka Abhijit IIHR, Bangalore
Arka Shreshtha IIHR, Bangalore
Arka Vardan IIHR, Bangalore
Arka Vishal IIHR, Bangalore
Arka Rakshak IIHR, Bangalore
Arka Ananya IIHR, Bangalore
Arka Samrat IIHR, Bangalore
Pusa Divya IARI, New Delhi
Pusa Hybrid-1 IARI, New Delhi
TH-802 PAU, Ludhiana
TH-2312 PAU, Ludhiana
RHRTH-57 MPKV, Rahuri
RHRTH-92 MPKV, Rahuri
COTH-1 TNAU, Coimbatore.
VRTH-1 IIVR,Varanasi
VRTH-2 IIVR,Varanasi
LIST OF F1 HYBRIDS IN
TOMATOHYBRID INSTITUTE
Rupali IAHS
Vaishali IAHS
Naveen IAHS
Rashmi IAHS
Megha Beejo Sheetal Seeds
Madhuri Beejo Sheetal Seeds
Manisha Beejo Sheetal Seeds
Meenakshi Beejo Sheetal Seeds
ARTH-3 Ankur Seeds
ARTH-4 Ankur Seeds

Arka Rakshak
• First public triple disease resistant tomato F1 hybrid in India
• High yielding F1 hybrid giving yield of 90-100 tons per hectare in 140-150
days
• Triple disease resistance to tomato leaf curl virus, bacterial wilt and early
blight
• Suitable for summer, kharif and rabi seasons
• Crossing an advanced breeding line bred at IIHR with another breeding line
bred at AVRDC, Taiwan

MAJOR CONSTRAINTS
 High cost of F1 hybrid seeds
 Lack of awareness among the growers about hybrid
crop production techniques
 Unorganized marketing system for vegetables
 Lack of postharvest management techniques
 Non availability of quality seeds
 Non availability of other inputs at proper time
 Non availability of biotic stress resistant hybrids

Advances in hybrid seed production of tomato

Advances in hybrid seed production of tomato

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