In Vitro Conservation And Cryopreservation In Tuber Crops
In Vitro Conservation andCryopreservation in Tuber CropsEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Root and tuber crops are second only in importanceto cereals as a global source of carbohydrates. They also provide some minerals and essentialvitamins. The variability of various root and tuber crops isdecreasing due to drastic changes in food habitscropping pattern, deforestationexcessive land utilizationextinction of landraces due to the overwhelmingpopularization of improved varietiesEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Root and Tuber Crops siaPapua New GuineaNambiaEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
India holds a rich genetic diversity of tropical root and tuber crops,namely, cassava, sweet potato, aroids, yams and several minortubers.There are five major areas of distribution of root andtuber crops in India.(i) South-western hilly and coastal region(ii) Southern peninsular region(iii) Eastern coastal region(iv) North-eastern region(v) North-western regionMaximum species diversity is noticed in North-easternand South-western hilly and coastal region.Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Important root and tuber crops in IndiaBotanical nameCommon name FamilyRegions of distributionManihot esculentaCrantzCassavaEuphorbiaceaeSouthern, occasionally ineastern regionsIpomoea batatas (L.)Lam.Sweet potatoConvolvulaceaeIntroduced,countryDioscorea alata L.Greater yamDioscoreaceaeSouthern and North-eastern regionsDioscorea rotundataPoir.White yamDioscoreaceaeIntroduced, Southern and Northeastern regionsDioscorea esculenta(Lour.) Burk.Lesser yamDioscoreaceaeSouthern, North-eastern and easternregionsDioscorea bulbifera L.var. sativaPotato yamDioscoreaceaeSouthern, North-eastern and easternregionsColocasia esculenta (L.) TaroSchott.AraceaeThroughout India with greaterdiversity in North-eastern, easternand southern regionXanthosomasagittifolium(L.) Schott.AraceaeSouthern and eastern IndiaTanniafoundallNorth-overEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGRthe
Important root and tuber crops in IndiaBotanical nameCommon name FamilyRegions of distributionAmorphophallus paeoniifolius(Dennst.) NicolsonElephantyamSouthern, North-easternand eastern regionsColeus parviflorus Benth.Pachyrhizus erosus (L.)UrbanChinese potatoLabiataeSouthern regionsYam beanFabaceaeNorth-eastern regionMoghania vestita Benth. exBak.MoghaniaFabaceaeNorth-eastern regionPsophocarpustetragonolobus (L.) DC.Winged beanFabaceaeSouthern and Northeastern regionsMaranta arundinacea L.ArrowrootMarantaceaeIntroduced, adapted toplains with high rainfallCanna edulis Ker-GawlerQueensland orCannaceaepurple arrowrootIntroduced, adapted to highrainfall regionsAlocasia macrorrhiza (L.) G.DonGiant taroSouthern and Northeastern regionsfoot AraceaeAraceaeEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Conservation of Tuber Crops Genetic Resource(National Status)ICAR-CTCRI, KeralaCropAccessions(Nos.)Cassava1211Sweet Potato1124Yams1110Edible Aroids683Minor Tuber crops200(Source: Annual report 2017-18)AICRP on Tuber CropsCropAccessions(Nos.)Cassava601Sweet Potato1927Taro733Yams453Edible Aroids430Minor Tuber crops438(Source: Annual report 2015-16)Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Conservation of Potato Genetic Resource(National Status)At ICAR-CPRI, Shimla, 4502 accessions of cultivated and wildpotato species, obtained from 30 countries, are conservedCropAccessions(Nos.)a) Tuberosum (Cultivars /parental lines)Cultivars bred at CPRI60Advanced hybrids96Indigenous varieties107Indigenous samples97Exotic2840b) Andigena762c) Wild/semi-cultivatedspp.540(125 species)(Source: CPRI Annual report 2018-19)Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Conservation of Tuber Crops Genetic Resource(ICAR-NBPGR Status)In field genebank, 283 accessions are being maintained at RS, ThrissurIn in vitro genebank, 518 accessions ( 7000 cultures) are being maintainedCrop spp.SpeciesAccession(No.)Cultures(No.)Sweet Potato(Ipomoeabatatas)12613000Dioscorea spp.101531850Taro 110100Alocasia indica1450Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
In vitro Culture Establishment in Sweet PotatoNodal explants areexcised, disinfectedand culturedCultures are established in 4-12weeks, depending on genotype,often requiring repeateddisinfectionCheck forContaminationDisinfectBacteria-free ClonesDiscardEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed/ refined for in vitro multiplication/ regenerationD. rotundataIpomoeabatatasDioscoreaalataD. bulbiferaD. pentaphyllaIn vitromultiplication/RegenerationD. floribundaD. nipponicaD. wallichiiD. AlocasiamacrorrhizaEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed for In vitroConservation of Tuber CropsNormal GrowthSlow GrowthD alataIpomoea batatasD bulbiferaD floribundaD pentaphyllaStorage Organs InductionD rotundataColocasia esculentaColocassia esculentaDioscorea spp.Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed for In vitroConservation of Tuber CropsNormal GrowthPlant speciesStrategyResponseReferenceD. alataMS 0.15mg/l NAA at 25 C10-12 monthsMandal et al.,2000D. bulbiferaMS 0.15mg/l NAA at 25 C10-12 monthsMandal et al.,2000D. pentaphylla MS 0.15mg/l NAA at 25 C10-12 monthsMandal et al.,2000D. rotundataMS 0.5 mg/l BAP 0.1 mg/l NAA at 10-12 months25 CMandal et al.,2000ColocasiaesculentaMS 0.5 mg/l BAP 0.1 mg/l NAA at 6-12 months25 CHussain et al.,2007aEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed for In vitroConservation of Tuber CropsSlow GrowthPlant speciesStrategyD. floribunda MS 0.5 mg/l NAA at 18 CResponseReference10-12 months Mandal et al.,2000IpomoeabatatasMS 0.2mg/l Kn 0.1mg/l IAA 16-18 months Mandal and4% sucrose (at 18 C)Chandel, 1991I. batatasMS 0.2mg/l Kn 0.1mg/l IAA 6-12 months4% sucrose (at 25 C)Mandal andChandel, 1991I. batatasMS 0.2mg/l Kn 0.1mg/l IAA 6-12 months2.0-2.5% sucrose (at 18 C)Mandal et al.,2000I. batatasMS 1.5- 2.0 % Mannitol at 25 C 6-12 monthsMandal et al.,2000Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
In vitro corm inductionExplant A shoot tip, embedded deep intothe corm tissue encircled with leafprimordial, and basal part of thepetioles (size: 5-10 mm diam, 2030 mm length)Media M1 MS (3% sucrose) 2.2 μM BAP 0.6μM NAA 0.8% agar M2 MS (8% sucrose) 2.2 μM BAP 0.6μM NAA 0.8% agar M3 MS (10% sucrose) 2.2 μM BAP 0.6μM NAA 0.8% agarGeneticStability Morphological Markers: 12 qualitativemorphological traits such as corm weight,corm bud colour, fresh weight of plant,corm length (cm), corm weight (g) no. ofeyes/corm Molecular markers: RAPD and ISSR markersEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed forCryopreservation of Tuber Crops4-6 week-old in vitro mother culturesShoot tips (1-2 mm)Pre-culture on MS medium having 0.3M sucroseEncapsulationdehydrationEncapsulation in 3% Calcium alginate &osmoprotection in liquid MS medium containing0.75 M sucrose for 1-4 daysAir dehydration in laminar flow for 5½ hrsPlunge in LNThawing at 40 C for 2 minRegrown on MS 1.5 mg/l BAP, 0.2 mg/l NAA and 0.2mg/l GA3Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Regeneration of encapsulated shoot tips of D. bulbifera after cryopreservationEffect of Sucrose ConcentrationSucroseConcentration (M)1Mukherjee et al. (2009) CryolettersPRECULTURE DURATION (Days)230.508.9 5.7 cd14.4 8.3 c17.8 7.9 bc0.7521.1 7.4 b43.3 8.2 a45.6 8.3 a1.04.4 4.9 de0.0 e0.0 eEffect of Preculture/ Dehydration DurationDehydrationDuration (hrs.)PRECULTURE DURATION (Days)12347414.4 8.335.6 9.638.9 7.437.8 6.330.0 6.7520.0 9.443.3 8.246.7 9.452.2 9.240.0 9.45½22.2 9.245.6 8.348.9 9.956.7 8.241.1 10.9620.0 9.437.8 7.940.0 9.447.8 9.236.7 13.3Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed forCryopreservation of Tuber CropsEncapsulation dehydrationPlant speciesCulturesystemResponseReference%survival% regenerationDioscorea alata Shoot tips7121Mandal et al., 1996D. rotundataShoot tips8867Mandal et al., 1999D. floribundaShoot tips7529Mandal, 2000;Mandal and AhujaGhosh, 2007D. wallichiiShoot tips6437Mandal et al., 1996IpomoeabatatasSomatic62% Regeneration of plants fromembryos /cryopreserved embryogenic tissueembryogenictissueMandal et al., 2000Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Vitrification4-6 week-old in vitro mother culturesShoot tips (1-2 mm)Pre-culture on MS medium having high sucroseTreat with loading solution (LS)PVS2 for 0-130 minPlunge in LNThawing at 40 C for 2 minUnloading solutionRegrowthFigure: Regrowth of Nodal explants ificationtechnique; a) Control, b) Vitrificationcontrol, c) Cryopreserved Nodal Explantsshowing regrowth, d) fully grown plantletfrom cryopreserved nodal explantEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Effect of exposure time to PVS2 at 0 C on survivaland regeneration of shoot tips of D. bulbiferaPVS2 ExposureTime (min)-LNSurvival(% SD) LNRegeneration(% SD)Survival(% SD)Regeneration(% SD)095.6 4.9 a81.1 5.7 a0.0 d0.0 d5092.2 6.3 ab72.2 6.3 b63.3 8.2 c27.8 7.9 c7091.1 7.4 ab65.6 6.8 c78.9 7.4 b44.4 8.3 b9090.0 6.7 b64.4 4.9 c88.9 8.7 a57.8 6.3 a11085.6 8.3 c58.9 5.7 d81.1 9.9 b45.6 8.3 b13081.1 7.4 d50.0 9.4 e74.4 10.6 b28.9 10.9 cMukherjee et al. (2009) CryolettersEighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
Protocols developed forCryopreservation of Tuber Crops(Vitrification Technique)Plant speciesPVS2 TreatmentDuration (min)ResponseReference% survival % regenerationDioscorea alata 90 min8239Mukherjee et al.,2009D. bulbifera90 min8958Mukherjee et al.,2009D. rotundata110 min8971Mandal et al.,2008D. wallichii90 min7128Mandal et al.2000Eighth International Training on In vitro and Cryopreservation Approaches for Conservation of PGR
TARO: FROM LAB TO LANDTARO INMEDIUM TERM CONSERVATIONAT IN VITRO GENE BANKESTABLISHMENT OF PLANTLETSAT RS- THRISSURKerala farmers’ request toDr. Joseph K. John regardingland races of taroSupplied 40 accessions ofColocasia esculenta(including landraces)PRIMARY HARDENINGAT NBPGRICAR-NBPGR Regional Station - Thrissur, Keralafor “On Farm Conservation”AT FARMER’S FIELD
Genetic diversity assessment D. alataIn vitro conserved D. alata accessions were analysed for geneticdiversity using genomic and EST-SSR markers.Dioscorea alata accessions IC044217 and IC-044214; IC044235AandIC-044233showed maximum similarity of98% .M123456789 10 11 12 13 14 15 16 17170150M181920212223242526272829500M 123456789 10 11 12 13 14 15 16 17260230200M181920212223242526272829
Genetic Diversity Analysis of Sweet Potato Genetic diversity analysis in in vitro conserved I batatas accessions wasdone using EST-SSR markers The accessions viz. EC-324082 (CIP, New Delhi), EC-64037 (NBPGR,Regional Station, Amravati), EC-13468 (NBPGR, Regional Station,Amravati) were found to be 4613EC-3393520.600.700.80Coefficient0.901.00
AcknowledgementsDr. B B MandalDr. Neelam SharmaDr. Zakir HussainDr. Vartika SrivastavaMr. Devendra K NerwalMr. Suresh Chand Mali
Root and tuber crops are second only in importance to cereals as a global source of carbohydrates. They also provide some minerals and essential vitamins. The variability of various root and tuber crops is decreasing due to drastic changes in food habits cropping pattern, deforestation excessive land utilization extinction of landraces due to the overwhelming
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The workshop included a session devoted to opportunities for the cryopreservation of aquatic species, in which presenters provided information about current efforts and approaches to preservation of aquatic model germplasm. Another session focused on current efforts addressing reproducibility and
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