ADVANCES IN HYBRID SEED ADVANCES IN HYBRID SEED PRODUCTION TECHNOLOGY OF TOMATO PRODUCTION TECHNOLOGY OF TOMATO Speaker: Akshay Chittora Ph.D. Horticulture
ADVANCES IN HYBRID SEED ADVANCES IN HYBRID SEED PRODUCTION TECHNOLOGY OF PRODUCTION TECHNOLOGY OF
TOMATOTOMATOSpeaker:
Akshay ChittoraPh.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
TOMATOTOMATOCommon 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 marketed in India by IAHS in 1973
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 linesIn Self-Pollinated : Pure lineIn Cross-Pollinated : Inbred (by selfing)
Testing of combining ability
GCA for additive Gene ActionsSCA for dominant Gene ActionPredictive Information from SCA by
(Single Cross, Double Cross, Three way Cross, Top Cross Poly Cross and Diallele)
Improvement of inbred lines / varietiesFor disease and quality trait
Production of hybrid seedTypes of Hybrids and their Seed ProductionHybridization
The commonly utilized mechanisms for developing
commercial hybrids in vegetablesMechanism 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 pollinationNP: Natural pollination
FLOWERING BEHAVIOURANTHESIS DEHISCENCE RECEPTIVITY
OF STIGMAStarts at 6 AM and maximum flower opening till late morning.
8 AM – 11 AM. 16 hrs before and 5 days after anthesis.
Hand emasculation and Hand emasculation and pollination as hybridization pollination 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 openP1: 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)
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 male Maintenance of male sterilitysterility
• Genic or pollen male sterilitymsms (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, 2006)
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 separatestamens and split pistils
Chandra Sekharand Sawhney,1990
Gibberlin synthesisCCC 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 commercialutilization
Moore, 1959
Dalapon Male sterility was induced Brauer, 1959TIBA (TriiodobenzoicAcid)
Some degree of male sterility wasinducing
Rehm, 1952
NAA Induced male sterility McRae, 1985
Biotechnological tools useful in Hybrid Biotechnological tools useful in Hybrid seed productionseed 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 haploid Application of haploid cultureculture
CROP FINDINGS REFERENCE
Tomato Haploidy has been successfully used for developing male sterile pure line
Zamit et al., 1980Schereva 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 malesterility
Monogenic recessive mutant was utilized todevelop cost effective experimental crosses.
Sawhney, 1997;Kumar et al., 2001
Auxotrophy A very attractive and feasible model wasproposed utilizing monogenic recessivenutritional mutants e.g. thiamin dependent.
Barabas, 1991
Incongruity
Models to transfer barrier (incompatibilitybetween pollen and stigma) genes andcorresponding penetration genes weredemonstrated.
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
Pusa Hybrid-2 IARI, New Delhi
Pusa Hybrid-4 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