Regeneration of transgenic cassava plants (Manihot esculenta Crantz) from microbombarded embryogenic suspension cultures

A protocol was established for the introduction of DNA into embryogenic suspension-derived tissues of cassava via microparticle bombardment, for the selection of genetically transformed cells, and for the regeneration of fully transgenic plants from these cells. The plasmid DNA used for bombardment contained a gene encoding neomycin phosphotransferase (nptII) and a gene encoding ß-glucuronidase (uidA). Selection of bombarded tissue with paromomycin resulted in the establishment of putative transgenic embryogenic calli. In most of these calli, ß-glucuronidase was detected histochemically. Molecular analysis of paromomycin-resistant embryogenic calli and of plants regenerated from these calli, confirmed the stable integration of bombarded DNA into the cassava genome.

[1]  Nigel J. Taylor,et al.  Development of friable embryogenic callus and embryogenic suspension culture systems in cassava (Manihot esculenta Crantz) , 1996, Nature Biotechnology.

[2]  J. Messing [2] New M13 vectors for cloning , 1983 .

[3]  B. Halkier,et al.  POSSIBLE USE OF A BIOTECHNOLOGICAL APPROACH TO OPTIMIZE AND REGULATE THE CONTENT AND DISTRIBUTION OF CYANOGENIC GLUCOSIDES IN CASSAVA TO INCREASE FOOD SAFETY. , 1994 .

[4]  A. W. Moore,et al.  Cassava viruses and genetic engineering , 1992 .

[5]  C. Fauquet,et al.  Transformation in Cassava (Manihot esculenta Crantz) , 1993 .

[6]  R. U. Schenk,et al.  Medium and techniques for induction and growth of monocotyledonous and dicotyledonous plant cell cultures , 1972 .

[7]  G. Henshaw,et al.  Somatic Embryogenesis in Cassava , 1982 .

[8]  Robert Kay,et al.  Duplication of CaMV 35S Promoter Sequences Creates a Strong Enhancer for Plant Genes , 1987, Science.

[9]  R. Beachy,et al.  A DNA sequence element that confers seed‐specific enhancement to a constitutive promoter , 1988, The EMBO journal.

[10]  J. Widholm The use of fluorescein diacetate and phenosafranine for determining viability of cultured plant cells. , 1972, Stain technology.

[11]  P. Shewry,et al.  Transient gene expression in cassava somatic embryos by tissue electroporation , 1995 .

[12]  W. Frommer,et al.  2 – The GUS Reporter System as a Tool to Study Plant Gene Expression , 1992 .

[13]  H. Klee,et al.  Rapid assay of foreign gene expression in leaf discs transformed by Agrobacterium tumefaciens: Role of T-DNA borders in the transfer process. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[14]  J. Cock Cassava: New Potential For A Neglected Crop , 1984 .

[15]  G. Hahne,et al.  Genotype and composition of culture medium are factors important in the selection for transformed sunflower (Helianthus annum)callus , 1991 .

[16]  F. Skoog,et al.  A revised medium for rapid growth and bio assays with tobacco tissue cultures , 1962 .