Research & Development

CRYOPRESERVATION

Until two decades ago the genetic resources were getting depleted owing to the continuous depredation by man. It was imperative therefore that many of the elite, economically important and endangered species are preserved to make them available when needed. The conventional methods of storage failed to prevent losses caused due to various reasons. A new methodology had to be devised for long-term preservation of material.

Cryopreservation is a non-lethal storage of biological material at ultra-low temperature. It is a process where tissues, organelles, cells, extracellular matrix, organs or any other biological constructs susceptible to damage caused by unregulated chemical kinetics are preserved by cooling to very low temp -196 degree Celsius using liquid nitrogen. At the temperature of liquid nitrogen (-196 degree) almost all metabolic activities of cells are ceased and the sample can then be preserved in such state for extended periods. However, only few biological materials can be frozen to (-196 degree) without affecting the cell viability.
Major advantages are: 1. once the material is successfully conserved to particular temperature it can be preserved indefinitely. 2. Once in storage no chance of new contamination of fungus or bacteria. 3. Minimal space required. 4. Minimal labor required. It is ideal method for long term conservation of material. Disease free plants can be conserved and propagated. Recalcitrant seeds can be maintained for long time. Endangered species can be maintained. Pollens can be maintained to increase longitivity. Rare germplasm and other genetic manipulations can be stored.

The expensive equipment needed to provide controlled and variable rates of cooling/warming temperatures can however be a limitation in the application of in vitro technology for large scale germplasm conservation. Formation of ice crystal inside the cell should be prevented as they cause injury to the cell. Sometimes certain solutes from the cell leak out during freezing. Cry protectant also effect the viability of cells.

In theory, all metabolic activities at temperatures of liquid nitrogen (LN) are reduced to zero, so that after rewarming from cryopreservation, true-to-type plants are expected. Since cryopreservation protocols do not only involve cooling in and rewarming from LN but also in vitro culture and regeneration processes, phenotypic and genomic changes can occur due to variation.

Therefore, the verification of true-to-type plants after cryopreservation is necessary.

Aqief Afzal
ASRBC, ACI LTD.