Slow growth storage of banana germplasm

Contributors to this page: Bioversity International, Belgium (Ines Van den Houwe); IITA, Nigeria (Dominique Dumet, Badara Gueye).

Sample processing
Inventory of stored cultures

Sample processing for tissue culture banks

Multiplication of propagules for conservation

When an accession has successfully passed the initiation phase (click here for details), then it is ready to be multiplied for storage, either for normal growth or slow growth conservation. This multiplication phase is also required for rapid propagation of selected materials for research or distribution.

Starting material

This propagation phase of an accession may vary from a few weeks to a few months as the multiplication rate strongly depends on the genomic group to which the accession belongs and is influenced by the composition of the medium (particularly the cytokinin concentration), the explant size, age of culture and the size of the culture vial.

Visual inspection of plant materials

Disposal of contaminated materials

Watch the video illustrating the aseptic subculturing process of banana shoots


Or view the video on Youtube

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Storage for tissue culture banks

Sample specifications

Container specifications

Growth media

Culture facility regimes

Storage duration

If the above physical and chemical storage conditions are followed, an average period of 12 months can be expected before re-culturing is required. These storage conditions are minimal growth conditions that proved to be acceptable for most genotypes. Not all accessions and genotypes, however, respond equally well to the applied conditions. (Further reading: Van den houwe et al. 1995).

System for tracking materials/inventory system during tissue culture storage

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Monitoring the performance of tissue cultures in slow growth storage

Need for subculturing multiplication of stored cultures

Quantitative and qualitative criteria should be considered to define the moment that an accession should be recycled after the material has been maintained for a given time in storage.

The spare cultures from the previous storage cycle can be discarded when the new set of cultures are transferred to the cold storage conditions described above.

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Inventory and monitoring of stored cultures

Accessions growing in tissue culture need to be inventoried and monitored on a timely basis in order to assess the number of healthy cultures remaining in storage and to determine the need for subculturing. The interval for routine visual monitoring of an active Musa collection should be about one month. Each individual culture in the accession tray should be checked visually and unsuitable cultures removed.

Checking the quality of the plantlets

The following factors should be taken into account to determine the quality of stored accessions:

Plantlets of unacceptable quality should be immediately discarded.

Checking the number of replicates

Monitoring genetic stability

Unless germplasm is regularly regenerated and transferred to the field for morphological observations, combined with the use of cytological techniques, genetic stability of a certain sample cannot be ascertained. Occasionally, abnormalities can be assessed in the in vitro samples.

In vitro assessment of variation

One of the criteria for efficient in vitro storage of germplasm is the maintenance of original genotypes over long periods of time. Although organized cultures (meristems, shoot and root-tip cultures) are believed to be genetically more stable than disorganized cultures (cell suspensions, protoplasts, callus, differentiated cells) variation appears to be relatively widespread in micro-propagated plants.

Factors like the culture mode, time in culture, number of subculture cycles, genotype and composition of the culture medium are known to influence the occurrence of somaclonal variation. The type and frequency of variation in micropropagated banana plants is known to be genotype and cultivar dependent.

Observations under greenhouse and field conditions (regeneration)

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References and further reading

Murashige T, Skoog F. 1962. A revised medium for rapid growth and bioassays with tobacco cell cultures. Physiologia Plantarum 15:473–497. Available for purchase here.

Strosse H, Van den houwe I, Panis B. 2004. Banana cell and tissue culture – review. Mohan Jain S, Swennen R (ed). Banana Improvement:Cellular, Molecular Biology, and Induced Mutations. Science Publishers Inc., Enfield, NH, USA:1-12.

Van den Houwe I, De Smet K, Tezenas du Montcel H, Swennen R. 1995. Variability in storage potential of banana shoot cultures under medium term storage conditions. Plant Cell, Tissue and Organ Culture 42:267-274. An abstract and full preview of the publication is available here.

Van den Houwe I, Panis B. 2000. In vitro conservation of banana: medium term storage and prospects for cryopreservation. Razdan MK, Cocking E, editors. Conservation of Plant Genetic Resources in vitro. Vol. 2. M/S Science Publishers, U.S.A. pp. 225-257.

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The Genebanks

The 11 CGIAR genebanks currently conserve 730,000 of cereals and grain legumes, forage crops, tree species, root and tuber crops, bananas and crop wild relatives.