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Application of tissue culture in crop improvement

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Application of tissue culture in crop improvement

  1. 1. Application of tissue culture in crop improvement Presented by, M. Uma Maheshwari, M. Phil., (Botany)
  2. 2. Introduction  Plant tissue culture comprises a set of invitro techniques, methods, strategies that are part of plant biotechnology.  Tissue culture had been exploited to create genetic variability, to improve the health of planted material, increase the number of desired germplasms.  Tissue culture are combined with molecular techniques, successfully used toincorporatespecifictraitsthroughgenetransfer.  Thecultureofprotoplasts,anthers,microspores,ovulesandembryos used tocreatenewgeneticvariation.  Cell culture produced somaclonal and gametoclonal variants with crop improvementpotential.  The culture of single cells and meristems can be effectively used to eradicatepathogenstherebyincreasingtheyieldofthecrops.
  3. 3. Application of tissue culture in crop improvement Newer molecular and cellular technologies have a broad significant impactoncropimprovement The various application of tissue culture approaches to crop improvement are- Breedingandbiotechnology Widehybridization Haploids Somaclonalvariation Micropropagation Syntheticseeds Pathogeneradication Germplasmpreservation
  4. 4. Breeding and biotechnology Plantbreedingseparatedintotwoactivities  Manipulatinggene  Plantevaluation Controlled pollination of plants lead to specific crosses result in new generation performedbetterintheyield Tissue–culture techniques is having significant impact in manipulating genetic diversity More than 50 different species have already been genetically modified either by vector dependent (Agrobacterium) or vector independent (Biolistic, micro- injectionandliposome)methods Tissue culture techniques have been played a major role in the development of plantgeneticengineering,especiallyinefficientgenetransferandtransgenicplant recovery
  5. 5. Wide hybridization Acriticalrequirementforcropimprovementistheintroductionofnew geneticmaterialintothecultivatedlinesofinterest,whetherviasingle genes, throughgeneticengineeringormultiplegenesthroughconventional hybridizationortissueculturetechniques Theprocessoffertilizationcanbeblockedatanyofthestages,resultingin thefunctionalbarriertothehybridizationandtheblockageofgenetransfer betweenthetwoplants. Twotypesof barriers Pre-zygoticbarriers Post-zygoticbarriers Toovercometheseproblems,thefollowingproceduresarefollowed invitrofertilization Embryoculture Protoplastfusion
  6. 6. in vitro fertilization  IVF has been used to facilitate both interspecific and intergeneric crosses, to overcome physiological basedselfcompatibilityandtoproducehybrids Awiderangeofplantshavebeenrecovered suchas  Tobacco  Clover  Corn  Rice  Canola  Cotton  Poppy
  7. 7. Embryo culture Itdealswiththesterileisolationandinvitrogrowthofamatureoranimmatureembryo ofobtainingaviableplant Seeddormancycanbesuccessfullybyepassed Twotypesofembryoculture Matureembryoculture Immatureembryoculture/Embryorescue Matureembryoculture o Matureembryosareisolatedfromripeseedsandare culturedin vitro o Itiscarriedoutduringfollowingcondition o Embryosremaindormantforlongtime o Lowsurvivalofembryosinvivo o Toavoidinhibitioninseedgermination o Convertsterileseedsintoviableseedlings
  8. 8. Embryo culture
  9. 9. It has been successfully applied to a number of agricultural crops including cotton, barley, banana, orchids, roses, tomato, rice, jute and some brassicas Application of embryo culture o Prevention of embryo abortion o Overcoming seed dormancy o Shortening of breeding cycle o Production of haploids o Overcoming seed sterility o Clonal propagation Embryo rescue o It involves the culture of immature embryos to rescue them from unripe or hybrid seeds that fails to germinate
  10. 10. Protoplast fusion It has been suggested as a means of developing unique hybrid plantswhichcannotbeproducedbyconventionalhybridization Protoplasts of two plants can be fused by chemical or physical methods Applicationofprotoplastfusion Ithas opened new possibilities for thein vitro genetic manipulation oftoimprovethecrops  Diseaseresistance  Environmentaltolerance  Qualitycharacters Cytoplasmicmalesterility
  11. 11. Protoplast fusion
  12. 12. Haploids  Haploid plants are characterized by possessing only a single set of chromosomes.  Haploid plants are of great significance for the production of homozygous lines  At least 171 plant species have been used to produce haploid plants by pollen,antherandmicrosporeculture  Cereals(barley,maize,rice,rye,wheat)  Fruits(grapeandstrawberry)  Oil-seeds(rapeandcanola)  Trees(apple,litchi,poplarandrubber)  Plantationcrops(cotton,sugarcaneandtobacco)  Vegetable crops (asparagus, Brussels sprouts, cabbage, carrot, pepper,potato,sugarbeet,tomatoandwingbean)
  13. 13. Haploids
  14. 14. Androgenesis  Haploidproductionoccurs throughantherorpollen culture  Intheplantbiotechnologyprogramme,haploidproductionisachievedby twomethods  Androgenesis  Gynogenesis Gynogenesis  Haploidproductionoccurs throughovaryorovule culture Pollenculturepreferablethanantherculturebecause • Androgenesis,startingfromasinglecellcanberegulated • Isolated micro spores are ideal for several genetic manipulations (transformation,mutagenesis) • Theyieldof haploidplantsisrelativelyhigher • Undesirableeffectofantherwallandassociatedtissuescanbeavoided
  15. 15. Somaclonal variation  The genetic variations found in the in vitro cultured cells are collectively calledsomaclonalvariations.Theplantsarecalledsomaclones  Itoccursduetogeneticheterogeneityinplanttissuecultures  Expressionofgeneticdisorders  Spontaneousmutationduetocultureconditions The geneticchangesinclude  Polyploidy  Aneuploidy  Chromosomal breakage  Deletion  Translocations  Geneamplification  Severalmutations
  16. 16.  Many of the changes observed in plants are regenerated in vitro have potential agriculturalandhorticulturalsignificance  Thealterationsinclude  Plantpigmentation  Seedyield  PlantvigourandSize  Essentialoils  Leafandflowermorphology  Fruitsolids  Diseasetoleranceorresistance Such variations are observed in wheat, maize, tomato, sugarcane,oats,oilseedrape,celery,potato
  17. 17. Somaclonal variation
  18. 18. Micro propagation invitroclonalpropagationthroughtissuecultureis calledmicropropagation Themicropropagationtechniqueispreferredto conventionalasexualpropagation Smallamountoftissueisneeded Possiblealternativefordevelopingresistance Meansforinternationalexchangeofplant materials invitrostockcanbeeasilyproliferated Micropropagationinvolvesinvitroclonalpropagationbytwo approaches  Multiplicationbyaxillarybuds/apicalbuds Multiplicationbyadventitiousroots
  19. 19. • Selection of mother plant and its maintenanceStage 0 • Initiation and establishment of cultureStage 1 • Multiplication of shoots or rapid somatic embryo formationStage 2 • in vitro germination of somatic embryos and or rooting of shootsStage 3 • Transfer of plantlets to sterilized soil for hardening under green house condition Stage 4 Major stages involved in Micro propagation
  20. 20. Micro propagation Applications  High rate of plant propagation  Production of disease-free plants  Production of seeds in some crops  Cost-effective process  Automated micro propagation Disadvantages x Contamination of cultures x Brewing of medium x Genetic variability x Vitrification x Cost factor
  21. 21. Synthetic seeds Usually, Sodium or Calcium alginate is selected for encapsulation becauseitislesstoxictoembryosandeasytohandle The artificial seeds can be maintained in a viable state still they are planted Asynthetic or artificial seed has been defined as a somatic embryo encapsulatedinsideacoatingandisconsideredtobeanalogoustoa zygoticseed Differenttypesofsyntheticseedsare Somaticembryosencapsulatedinawatergel Dried andcoatedsomaticembryos Driedanduncoatedsomaticembryos Somaticembryosuspendedinafluidcarrier Shootbudsencapsulatedinawatergel
  22. 22. Pathogen eradication Plant species are infected with pathogens-viruses, bacteria, fungi, mycoplasma and nematodesthatcausesdiseases Tissue culture techniques employing meristem-tips are successfully used for the productionofdisease-freeplants List of plants with virus-elimination by meristem-culture Plant species Virus eliminated Solanum tuberosum Leaf roll potato virus-A, X, Y, S Allium sativum Mosaic virus Petunia spp. Tobacco mosaic virus Musa spp. Cucumber mosaic virus Brassica oleracea Cauliflower / turnip mosaic virus Armoracia rusticena Turnip mosaic virus
  23. 23. Germplasm preservation Germplasm broadly refers to the hereditary material transmittedtotheoffspringthroughgermcells The main objective of germplasm conservation is to preserve the genetic diversity of a particular plant or geneticstockforitsuseatanytimeinfuture in vitro methods employing shoots, meristems and embryosareideallysuitedforgermplasmconservation Threemainapproaches  Cryopreservation  Coldstorage  Low-pressure and Low-oxygen storage
  24. 24. Advantages  Largequantitiescanbestoredinasmallarea Maintained in an environment free from pathogens Protectedagainstnature’shazards From germplasm stock, large number of plantscanbeobtainedwheneveritisneeded Obstacles for their transport through national and international borders are minimal (since theyaremaintainedunderasepticcondition)

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