In vitro Preservation of Germplasm SMG
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This document discusses various methods for in vitro preservation of germplasm, including cryopreservation, slow growth cultures, DNA clones, and desiccated somatic embryos. Cryopreservation involves freezing plant materials like shoot tips or cell cultures in liquid nitrogen at -196°C. It requires preculturing materials in cryoprotectants and slow cooling to prevent ice crystal formation. Slow growth cultures provide long-term storage at reduced temperatures or osmoticum concentrations with minimal subculturing. DNA clones allow conservation of genes by cloning DNA segments in vectors, while desiccated somatic embryos can be dried and stored at low temperatures like seeds. These methods help preserve crop diversity and valuable genes.
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This document provides information on in vitro germplasm conservation. It discusses that germplasm conservation aims to preserve the genetic diversity of plants. There are several methods of in vitro conservation including cryopreservation, cold storage, and low pressure/low oxygen storage. Cryopreservation involves freezing plant cells and tissues at ultra-low temperatures like in liquid nitrogen to bring their metabolism to zero. It allows for long term conservation of large amounts of genetic material in a small space. Cold storage conserves germplasm at low non-freezing temperatures to slow growth. Low pressure and low oxygen storage reduce atmospheric pressure and oxygen concentration to inhibit plant tissue growth.
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This document discusses techniques for isolating pure cultures of microorganisms and methods for preserving cultures. It describes various isolation techniques including streak plating, pour plating, spread plating, and using a micromanipulator. Methods for preserving cultures long-term include periodic transfer to fresh media, storage at low temperatures including liquid nitrogen, storage in sterile soil, overlaying with mineral oil, and lyophilization/freeze drying. Lyophilization is highlighted as one of the best methods as it allows viability for over 30 years without contamination and the cultures can be easily revived.
Preservation of pure culture.pptx
According to Lokesh, the goal of pure culture preservation is to preserve genetic stability and microbiological viability by use of several methods. Important techniques include the straightforward and inexpensive short-term storage of refrigeration at 4°C on agar slants; the long-term preservation of deep freezing at -70°C to -80°C with cryoprotectants like glycerol or DMSO, which guarantees little genetic changes over years; and the freeze-drying process, or lyophilization, in which cultures are frozen and dehydrated under a vacuum to preserve viability and stability over long periods of time. Research, clinical diagnostics, and industrial applications all depend on these techniques to guarantee the availability of clean microbial cultures when required.
preservation of microorganism
This document discusses various methods for preserving microorganisms in pure culture, including periodic transfer to fresh media, lyophilization (freeze drying), cryopreservation, storage under oil or in saline suspension, and drying in vacuum. The key objectives of preservation are to maintain isolated pure cultures for extended periods while avoiding contamination or genetic changes. Lyophilization and cryopreservation in liquid nitrogen are commonly used techniques that allow long-term viable storage for decades by stopping microbial growth and metabolism. Periodic transfer risks contamination and genetic variation over time.
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Conservation of plant genetic resources and cryopreservation
Conservation of plant genetic resources and cryopreservation
pureculturepreservation-meghali-181227111743 (1).pdf
pureculturepreservation-meghali-181227111743 (1).pdf
Germplasm Conservation || Presented by Mamoona Ghaffar
Germplasm Conservation || Presented by Mamoona Ghaffar
Germplasm Conservation || Presented by Mamoona Ghaffar
Germplasm Conservation || Presented by Mamoona Ghaffar
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In vitro Preservation of Germplasm SMG
- 2. In Vitro Preservation of Germplasm Germplasm – sum total of all the gens present in a crop and its related species. Represented by collections of various strains and species. Provide genes for crop improvement Conventionally stored as seed at ambient, low or very low temperature Some crops produce recalcitrant (short – lived seeds).
- 3. 1. Cryopreservation (Freeze Preservation) Preservation of cells, tissues and organs in liquid Nitrogen. Used for Germplasm conservation Tissue cultures may be frozen and stored in liquid Nitrogen at – 1960C Useful for root and tuber crops Meristematic cells survive more than mature cells.
- 4. Steps I. Freezing Material is partially dehydrated – increases the survival of cells or tissues (In vacuum or treating with plant vitrification solution – a specially formulated concentrated solution) A cryprotectant is added to the medium – prevents the formation of ice crystals in cells and tissues and protect from toxic solution effect due to water loss from the cytoplasm. Eg. Dimethyl Sulphoxide (DMSO, 5-8 %)
- 5. • Added gradually over a period of 30- 60 minutes and the temperature of the culture is maintained at 0oC Glycerol, Proline – 10% etc. can also be used. Cells and tissues are encaspsulated in a matrix – e.g. alginate Cooling is done slowly or rapidly or it may be initially slow and then cooled rapidly by plunging the vials into liquid Nitrogen.
- 6. II. Storage Frozen cells and tissues are stored in a liquid Nitrogen refrigerator. III. Thawing Vials are plunged into water at 37-400 C for 90 minutes. Transferred into an ice bath Reculture on a filter paper disc in a suitable culture medium. Frequently transfer into fresh medium The cells are scrapped off from the filtered disc and cultured directly on the medium.
- 7. IV. Reculture Reculture is done in a suitable medium. Requirements of culture medium vary for different plants (eg. Shoot tips from freeze – preserved seedling of tomato need GA3 (Gibberellic acid) for developing into shoots.
- 8. Applications: Used for cryopreservation of a number of species . Materials used : Cell cultures Shoot tips Somatic embryos Zygotic embryos. Plant lets.
- 9. 2. Slow growth cultures Simple, cheap and effective method Methods: o Maintain the plant lets at a low temperature 4- 9OC or 15OC or in a medium with high osmotic concentration (or high osmoticum – 20% Sorbitol or Sucrose). o Nutrient restriction
- 10. o Use of growth retardants • Use of culture vessels having longer gas diffusion distances (the distance from medium surface to the cap of culture vessels) – Plant lets cultured in such vessels in an autotrophic state require only inorganic nutrition. Eg:- In carrot, growth was reduced to 50% when the gas diffusion distance was increased to 150 mm from 50mm • Attention is necessary only once in several months. • Subculture may be needed only after very long periods.
- 11. Uses Germplasm conservation of root, tuber and tree species by NBPGR, New Delhi. Ginger Garlic Banana Sweet Potato
- 12. 3. DNA Clones o Germplasm conservation in the form of DNA segments cloned in a suitable vector(eg. Cosmids). o Used for the conservation of valuable genes or DNA segments from threatened species. o Expensive method.
- 13. 4. Desiccated somatic embryos and Artificial seeds • can be stored at low (40C) or very low (- 200C) temperatures for long periods. •Applicable only in cases of species where adequate regeneration of somatic embryo occur.
- 14. THANK YOU