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Plant tissue culture techniques of Banana

Commercial cultivation of banana using plant tissue culture technique.This provides a good job opportunity and a good business.......

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Plant tissue culture techniques of Banana

  1. 1. Banana Culture<br />R.Sathes<br />Student<br />Department of Plant Biology and Bio technology<br />Loyola College<br />
  2. 2. What is Tissue Culture?<br />Tissue culture is collection experimental methods of isolation and inoculation of organs, tissues and cell in an artificial medium under in-vitro aseptic condition.<br />Micro propagation is vegetative propagation of plant using plant tissue culture. This also known as direct differentiation.<br />
  3. 3. Banana Cultivation in India<br />Banana is a globally important fruit crop with 97.5 million tones of production. <br />In India it supports livelihood of million of people. <br />With total annual production of 16.91 million tones from 490.70 thousand ha., with national average of 33.5 T/ha.<br />Maharashtra ranks first in production with 60 T/ha. Banana contributes 37% to total fruit production in India.<br />Banana occupy 20% area among the total area under crop in India. <br />Jalgaon is a major Banana growing district in Maharashtra which occupy 50,000 hectares area under Banana. <br />But most of Banana is grown by planting suckers. <br />The technology development in agriculture isvery fast, it results in developing Tissue Culture Technique.<br />
  4. 4. Requirement for Banana Growth<br />
  5. 5. Agro Climate<br />Banana is basically a tropical crop, grows well in temperature range of 13ºC – 38ºC with RH regime of 75-85%. <br />In India this crop is being cultivated in climate ranging from humid tropical to dry mild subtropics through selection of appropriate varieties like Grandnaine. <br />The normal growth of the banana begins at 18ºC, reaches optimum at 27ºC, then declines and comes to a halt at 38ºC. <br />Higher temperature causes sun scorching. <br />High velocity wind which exceeds 80 km phrs damages the crop.<br />
  6. 6. Soil<br />Soil for banana should have good drainage, adequate fertility and moisture. Deep, rich loamy soil with pH between 6-7.5 are most preferred for banana cultivation. <br />Ill drained, poorly aerated and nutritionally deficient soils are not suitable for banana.<br />Saline solid, calcareous soil are not suitable for Banana cultivation. <br />Avoided soil of low laying areas, very sandy & heavy black cotton with ill drainage.<br />A soil that is not too acidic & not too alkaline, rich in organic material with high nitrogen content, adequate phosphorus level and plenty of potash are good for banana.<br />
  7. 7. Varieties<br />In India banana is grown under diverse conditions and production systems. Selection of varieties, therefore is based on a large number of varieties catering to various kinds of needs and situations.<br />Dwarf Cavendish.<br />Robusta. <br />Monthan<br />Poovan<br />Nendran<br />Red banana<br />Nyali<br />SafedVelchi<br />Basarai<br />Ardhapuri<br />Rasthali<br />Karpurvalli<br />Karthali <br />Grandnaine <br />Grandnaine is gaining popularity and may soon be the most preferred variety due to its tolerance to biotic stresses and good quality bunches. <br />
  8. 8. Grandnaine variety<br />
  9. 9. Planting Material<br />Sword suckers weighing approximately 500-1000 gm are commonly used as propagating material. <br />
  10. 10.
  11. 11. Why tissue culture?<br />Suckers generally may be infected with some pathogens and nematodes. <br />Similarly due to the variation in age and size of sucker the crop is not uniform, harvesting is prolonged and management becomes difficult.<br />Therefore, in-vitro colonel propagation i.e. Tissue culture plants are recommended for planting. <br />They are healthy, disease free, uniform and authentic. <br />Properly hardened secondary seedlings are only recommended for planting<br />
  12. 12. Advantages of Tissue Culture Planting Material<br />True to the type of mother plant under well management.<br />Pest and disease free seedlings.<br />Uniform growth, increases yield.<br />Early maturity of crop - maximum land use is possible in low land holding country like India.<br />Round the year planting possible as seedlings are made available throughout the year.<br />Two successive ratoons are possible in a short duration which minimizes cost of cultivation.<br />No staggered harvesting.<br />95% - 98% plants bear bunches.<br />New varieties can be introduced and multiplied in a short duration.<br />
  13. 13.
  14. 14. Micropropagation<br />Of Banana<br />
  15. 15. Initiation of shoot cultures<br />Shoot cultures of banana start conventionally from any plant part that contains a shoot meristem, i.e. the parental pseudo stem, small suckers, peepers and lateral.<br />For rapid in vitro multiplication of banana, shoot tips from young suckers of 40-100 cm height are most commonly used as explants. From the selected sucker a cube of tissue of about 1-2 cm³ containing the apical meristem is excised. <br />
  16. 16. The optimal size of the explants depends on the purpose. <br />For rapid multiplication, a relatively larger explants (3-10 mm) is desirable despite its higher susceptibility to blackening and contamination. <br />When virus or bacteria elimination is needed, meristem-tip culture is the preferred option. <br />The explants is then further reduced in size (0.5-1 mm length), leaving a meristematic dome with one or two leaf initials. <br />Meristem cultures have the disadvantage that they may have a higher mortality rate and an initial slower growth.<br />
  17. 17. The suckers for culture is prepared and taken into the lab.<br />They are soaked in Bavistin for 18 hours in order to remove the fungus and fungal spores.<br />In lab first they are washed in running water.<br />Then they are Dipped into detergent water (teepol) containing for one hour.<br /> After that they are taken and washed in tap water.<br />Washed suckers are taken to LAF chamber and further sterilization is done.<br />
  18. 18. In LAF chamber the suckers are sterilized using mercury chloride<br />Two different concentrations are used for sterilization purpose.<br />First the sucker sterilized using 0.12% of HgCl2. The sucker put into the bottle containing HgCl2 Shake well for 2 min.<br />After that HgCl2is removed and the sucker washed using distilled water. At first the distilled water added and the bottle shaked for 1 min. then the water removed and fresh distilled water added shaked for another one min. The water removed.<br />This step repeated under following timings 2min,3min,5min and 12 min.<br />
  19. 19. After 1st sterilization a layer of the sucker removed carefully.<br />The suckers are again sterilized using 0.1% HgCl2 for 5 min.<br />After that they are washed thoroughly with distilled water in following timings 1min,1min,2min,3min,5min,12min. <br />Finishing the above process another layer of sucker is removed.<br />The suckers are ready for inoculation.<br />
  20. 20. Suckers<br />Layer removal<br />1st Layer<br />2nd Layer<br />Inner Layer<br />2nd sterilization<br />1st sterilization<br />Removal of a layer<br />Removal of a layer<br />
  21. 21. Sterilization process<br />
  22. 22. Medium used for Banana cultivation<br />For banana micro propagation, MS-based media are widely adopted. Generally, they are supplemented with sucrose as a carbon source at a concentration of 30-40 g/l. Media are poured in a glass bottle where suckers are propagated.<br />Usually two types of growth regulators used, a cytokinin and an auxin, are added to the banana growth medium. <br />Their concentration and ratio determines the growth and morphogenesis of the banana tissue<br />In most banana micro propagation systems, semi-solid media are used. As a gelling agent agar (5-8 g/l) is frequently added to the culture medium.<br />
  23. 23. Problem of banana propagation<br />Banana tissue cultures often suffer from excessive blackening caused by oxidation of polyphenolic compounds released from wounded tissues. <br />These undesirable exudates form a barrier round the tissue, preventing nutrient uptake and hindering growth. <br />Therefore, during the first 4-6 weeks, fresh shoot-tips are transferred to new medium every 1-2 weeks. Alternatively, freshly initiated cultures can be kept in complete darkness for one week.<br /> Antioxidants, such as ascorbic acid or citric acid in concentrations ranging from 10-150 mg/l, are added to the growth medium to reduce blackening, or the explants are dipped in antioxidant solution (cysteine 50 mg/l) prior to their transfer to culture medium.<br />
  24. 24. Transfer to Growth room <br />Banana shoot-tip cultures are incubated at<br />An optimal growth temperature of 28 ± 2°C <br />In a light cycle of 12-16 h with a photosynthetic photon flux (PPF) of about 60 µE/m2s1.<br />Air condition will be working all the time to provide needed temperature. Which also provide clean dust free environment.<br />
  25. 25. Growth Room<br />
  26. 26. Inoculate sucker & Sucker after 2 weeks<br />
  27. 27. Arise of shoots from suckers<br />
  28. 28. Sub Culturing of Banana<br />After 2 weeks the suckers will become greenish in colour.<br />2 weeks after that multiple shoot will arise from the base of the suckers.<br />The shoots are cut at the base separated and placed in a fresh medium.<br />In each bottle three shoots are inoculated.<br />After a week multiple shoots arise from the inoculated shoot. Again they are separated and placed in a new fresh medium. <br />
  29. 29. Sub Cultured Banana<br />
  30. 30. Sub culturing and inoculated cultures<br />
  31. 31. The sub culturing is done until the require amount of plant needed.<br />The shoots are every day checked for contamination and the contaminated shoots are transferred to a fresh medium.<br />Meanwhile a set of well grown healthy shoots are taken for rooting.<br />
  32. 32. Well grown healthy shoots<br />
  33. 33. Rooting of Banana Shoots<br />Rooting of banana shoot is done in bottle containing charcoal medium.<br />For rooting IAA Used as growth regulator (for commercial purpose).<br />Medium without hormone gives good results.<br />It will take 2 weeks for rooting and fresh roots are arise at the base of the shoot.<br />
  34. 34. Rooted Plantlet<br />
  35. 35. Different Stages Of Banana Culture<br />
  36. 36. Acclimatization of banana plantlet <br />Rooted plantlet are kept in basal medium for certain time.<br />Then there are taken for acclimatization process where they gradually trained to the in-vivo temperature and light.<br />There are two stages in hardening process<br />Primary Hardening<br />Secondary Hardening<br />The hardened plantlets are carefully maintained in green house.<br />
  37. 37. Plates used for primary Hardening<br />
  38. 38. Secondary Hardening<br />
  39. 39. Transporting the plantlets.<br />The plantlets after acclamatization should be transported to the required place. Normal transportation is done where the plants are placed and grown in Plastic Bags.<br />Hence the well grown plants removed to provide the space in green house for the next cycle of plants and also to lower the cost of storage.<br />
  40. 40. Scope of banana Culture<br />Banana have lot of useful properties which built the pathway for banana growers.<br />Normal banana suckers which obtained directly from the plantation grows in 12 to 15 month where PTC plantlet will grow in 10 months.<br />The suckers are collected for PTC are healthy and high yielding so the daughter plantlets have the same character of the mother plant.<br />The banana is a short term crop so it provides immense opportunity to produce PTC plants through out the year.<br />Care full measures should be taken in growing PTC banana which can give a good job opportunity and income for the producers.<br />
  41. 41. Entrepreneurship is living a few years of your life like most people won’t, so that you can spend the rest of your life like most people can’t.<br />YOU CAN WIN<br />
  42. 42. Thank You<br />