Cryopreservation Studies in Cassava Tissues'

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Cryopreservation Studies in Cassava Tissues.

• R. Escobar, N. Manrique, F. Gil, L. Santos, L.
  Muñoz, D. Debouck, J. Tohme and W. Roca
• Centro Internacional de Agricultura Tropical CIAT
• Cali-Colombia

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Tissues and methods uses on cryopreservation

• Meristems
• Shoots
• Buds
• Seeds and embryos
• FEC
• Polen
• Somatic embryos

• Slow freezing , Rapid freezing
• Slow freezing , Rapid freezing,
• Encapsulation-dehydration , Vitrification,
• Encapsulation-vitrification
• Encapsulation-dehydration
• Slow freezing , Rapid freezing
• Encapsulation-dehydration, Vitrification,
• Desiccation, Desiccation-vitrification
• Slow freezing , Rapid freezing
• Slow freezing , Rapid freezing

CIAT, 1988-2001
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Escobar et al 1997
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Survival of cryopreserved shoot tips
shoot recovery
Significant differences (?0.05 level)
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Pre-culture medium and shoot recovery of
recalcitrant cv
shoot recovery
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Effect of cytokinins on plant recovery after
freezing (cv MCol 22)
shoot formation
Concentration mg/l
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Dehydration curve of cassava beads
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Encapsulation dehydration methodology
BRU, 2001
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Groups of response after freezing based on shoot
formation percentage
BRU, 2001
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Reproducibility of the response of six cassava
clones cryopreserved in L.N. across time
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Response of some clones cryopreserved during
different periods
BRU, 2001
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Field testing of cryopreserved clones
CIAT, Nov 1, 2001
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Response after freezing of different types of
explants
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Wild species tested with encapsulation
dehydration methodology
M. carthaginensis 413-013 M. carthaginensis
17F-417-001 M. esc. subs. peruviana 413-003 M.
esc. subs. peruviana 417-003 M. esc. subs.
peruviana 417-005 M. esc. subs. flabellifolia
443-001 M. esc. subs. flabellifolia 444-002 M.
esc. subs. flabellifolia 433-002
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Wild species response to desiccation time
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Cryopreservation of seeds and zygotic embryos
Marin et al. 1990 Survival after freezing steps
? 97 Slow and rapid method could be
applied Slow thawing (1h/-70C, 1 h/-15, room
temp) Adjustment protocol (BRU)
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Cryopreservation of FEC to aid transformation

• 4 FEC lines (4 cultivars) established at CIAT
• 8-12 month to establish FEC
• Instability and low regeneration rates with time
• M Nig 11 (TMS 60444) as control cultivar
• M Col 2215 and CM3306-4 as target clones
• Vitrification per se did not work (PVS2, PVS3)
• Desiccation worked with two cultivars
• Low cost, simple method for FEC bank

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Conclusions

• Cryopreservation plays a critical role in the
  integral, complementary, conservation strategy
  for cassava
• We developed 3 cryo. methodologies for cassava.
  From all of them we recovered plants, but
  varietal differences were observed.
• E-D seems to be the fastest and cheapest method
  to introduce the collection into cryo adjusted
  methods could be uses with wild species
• Dessication and D-V could be used as potential
  methods for cryo of FEC. It was possible to
  restore cell growth after freezing of FEC.
• A pilot project using the core collection will be
  strategic to establish logistic aspect in cryo.

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Acknowledgments
Maria Luisa Marin Guillermo Reina Mauricio
Giraldo Juan Diego Palacio M.P. Rangel Pablo
Herrera Miryam Cristina Duque Gerardo
Gallego IPGRI