Plant Molecular Biology Reporter

, Volume 19, Issue 4, pp 307–319 | Cite as

Phosphomannose isomerase: A versatile selectable marker forArabidopsis thaliana germ-line transformation

  • Rebecca Todd
  • Brian W. TagueEmail author


A new selection system using mannose has been evaluated for germ-line transformation ofArabidopsis thaliana. Although mannose itself has no adverse effects on plant cells, it leads to an accumulation of mannose-6-phosphate, which depletes intracellular stores of inorganic phosphate. This results in an inhibition of plant cell growth. The selection system uses theEscherichia coli pmi gene that encodes phosphomannose isomerase (PMI). Transgenic plants carrying thepmi gene can detoxify mannose-6-phosphate by conversion to fructose-6-phosphate, an intermediate of glycolysis, via the PMI activity. Germ-line transformation ofA. thaliana followed by sterile selection on 2–5 mM of mannose resulted in the isolation of mannose-6-phosphate-resistant progeny in about 2.5% of the treated seed, consistent with transformation rates using other selection schemes. Integrative transformation was confirmed by Southern hybridization. Analysis of PMI enzyme activity demonstrated a 5-fold range of activity levels, although these differences had little effect on the ability to select transformed plants or on the growth of transformed plants on mannose. Finally, mannose selection using thepmi gene could be accomplished in sterile plates and in soil, making this an extremely versatile tool forA. thaliana transformation.

Key words

germ-line transformation Arabidopsis thaliana mannose mannose-6-phosphate resistance phosphomannose isomerase 



phosphomannose isomerase


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  1. Altmann T, Damm B, Halfter U, Willmitzer L, and Morric P-C (1992) Protoplast transformation and methods to create specific mutants inArabidopsis thaliana. In: Koncz C, Chua N-H, and Schell J (eds), Methods in Arabidopsis Research, pp 310–330, World Scientific Publishing Co., River Edge NJ.Google Scholar
  2. Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, and Struhl K (1995) Current Protocols in Molecular Biology, 3rd edition. John Wiley & Sons, New York, NY.Google Scholar
  3. Baskin T, Remillong E, and Wilson J (2001) The impact of mannose and other carbon sources on the elongation and diameter of the primary root ofArabidopsis thaliana. Austral J Plant Phys 28: 481–488.Google Scholar
  4. Church GM and Gilbert W (1984) Genomic sequencing. Proc Natl Acad Sci (USA) 81: 1991–1995.CrossRefGoogle Scholar
  5. Clough SJ and Bent AF (1998) Floral dip: a simplified method forAgrobacterium-mediated transformation ofArabidopsis thaliana. Plant J 16: 735–743.PubMedCrossRefGoogle Scholar
  6. Ferguson J, Street H, and David SD (1958) The carbohydrate nutrition of tomato roots IV. The inhibition of excised root growth by galactose and mannose and its reversal by dextrose and xylose. Ann Bot 22: 525–538.Google Scholar
  7. Gill JF, Deretic V, and Chakrabarty AM (1986) Overproduction and assay ofPseudomonas aeruginosa phosphomannose isomerase. J Bacteriol 167: 611–615.PubMedGoogle Scholar
  8. Haldrup A, Petersen S, and Okkels F (1998) Positive selection: a plant selection principle based on xylose isomerase, an enzyme used in the food industry. Plant Cell Rep 18: 76–81.CrossRefGoogle Scholar
  9. Hansen G and Wright MS (1999) Recent advances in the transformation of plants. Trends Plant Sci 4: 226–231.PubMedCrossRefGoogle Scholar
  10. Joersbo M (2001) Advances in the selection of transgenic plants using non-antibiotic marker genes. Physiol Plant 111: 269–272.PubMedCrossRefGoogle Scholar
  11. Joersbo M, Donaldson I, Kreiberg J, Petersen SG, Brunstedt J, and Okkels FT (1998) Analysis of mannose selection used for transformation of sugar beet. Mol Breeding 4: 111–117.CrossRefGoogle Scholar
  12. Joersbo M and Okkels T (1996) A novel principle for selection of transgenic plant cells: positive selection. Plant Cell Rep 16: 219–221.CrossRefGoogle Scholar
  13. Lindsey K and Gallois P (1990) Transformation of sugar beet (Beta vulgaris) byAgrobacterium tumefaciens. J Exp Bot 41: 529–536.CrossRefGoogle Scholar
  14. Lowry OH, Rosebrough NJ, Farr AL, and Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–275.PubMedGoogle Scholar
  15. Lucca P, Ye X, and Potrykus I (2001) Effective selection and regeneration of transgenic rice plants with mannose as selective agent. Mol Breeding 7: 43–49.CrossRefGoogle Scholar
  16. Malca I, Endo R, and Long M (1967) Mechanism of glucose counteraction of inhibition of root elongation by galactose, mannose and glucosamine. Phytopathology 57: 272–278.Google Scholar
  17. Negrotto D, Jolley M, Beer S, Wenck AR, and Hansen G (2000) The use of phosphomannose-isomerase as a selectable marker to recover transgenic maize plants (Zea mays L.) via transformation. Plant Cell Rep 19: 798–803.CrossRefGoogle Scholar
  18. Pego JV, Weisbeek PJ, and Smeekens SCM (1999) Mannose inhibits arabidopsis germination via a hexokinase-mediated step. Plant Physiol 119: 1017–1023.PubMedCrossRefGoogle Scholar
  19. Sigma-Aldrich Fine Chemicals (2001) Biochemicals and Reagents for Life Science Research. Sigma-Aldrich, St Louis MO.Google Scholar
  20. Vergunst AC, de Waal EC, and Hooykaas PJJ (1998) Root transformation byAgrobacterium tumefaciens. In: Martinez-Zapater J and Salinas J (eds), Arabidopsis Protocols, pp 227–244, Humana Press Inc., Totowa NJ.CrossRefGoogle Scholar
  21. Wang AS, Evans RA, Altendorf PR, Hanten JA, Doyle MC, and Rosichan JL (2000) A mannose selection system for production of fertile transgenic maize plants from protoplasts. Plant Cell Rep 19: 654–660.CrossRefGoogle Scholar
  22. Zhang P, Potrykus I, and Puonti-Kaerlas J (2000) Efficient production of transgenic cassava using negative and positive selection. Transgenic Res 9: 405–415.PubMedCrossRefGoogle Scholar
  23. Zhang P and Puonti-Kaerlas J (2000) PEG-mediated cassava transformation using positive and negative selection. Plant Cell Rep 19: 1041–1048.CrossRefGoogle Scholar

Copyright information

© International Society for Plant Molecular Biology 2001

Authors and Affiliations

  1. 1.Department of BiologyWake Forest UniversityWinston-SalemUSA

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