Abstract
The predominant storage carbohydrates of mature carrot (Daucus carota L.) storage roots typically are the free sugars glucose and fructose. This trait is conditioned by the Rs allele. A naturally occurring recessive mutation, rs/rs, conditions a shift from these reducing sugars to sucrose. RT-PCR and sequencing revealed a unique 2.5 kb insert in the first and largest intron near the 5′ end of the acid soluble invertase isozyme II gene of rs/rs carrots. This insert was not totally spliced out during mRNA processing. While the wild-type acid-soluble invertase isozyme II transcript (ca. 2 kb) was detected in Rs/Rs roots and leaves, none was observed in rs/rs roots throughout development. RT-PCR of rs/rs leaves revealed two novel transcripts (2.7 kb and 3.2 kb). A comparison of enzyme activity between the near-isogenic Rs/Rs and rs/rs carrot lines revealed very low acid-soluble invertase activity in rs/rs roots whereas neutral invertase, sucrose synthase and sucrose phosphate synthase levels were comparable. Those results and linkage analysis indicate that Rs is a candidate locus for carrot vacuolar acid-soluble invertase isozyme II. Although the 2.5 kb insert does not occur in the Rs wild-type acid-soluble invertase isozyme II allele, it does occur elsewhere in the genome of Rs/Rs plants.
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References
Alabran, D.M. and Mabrouk, A.F. 1973. Carrot flavor, sugars, and free nitrogenous compounds in fresh carrots. J. Agric. Food Chem. 21: 205–208.
Casa, A.M., Brouwer, C., Nagel, A., Wang, L., Zhang, Q., Kresovich, S. and Wessler, S.R. 2000. The MITE family Heartbreaker (Hbr): molecular markers in maize. Proc. Natl. Acad. Sci. USA 97: 10083–10089.
Cheng, W.H., Taliercio, E.W. and Chourey, P.S. 1996. The Miniature 1 seed locus of maize encodes a cell-wall invertase required for normal development of endosperm and maternal cells in the pedicel. Plant Cell 8: 971–983.
Copeland, L. 1990. Enzymes of sucrose metabolism. In: P.J. Lea (Ed.) Methods in Plant Biochemistry: Enzymes of Primary Metabolism Vol. 3, Academic Press, New York, pp. 73–84.
Freeman, R.E. and Simon, P.W. 1983. Evidence for simple genetic control of sugar type in carrot (Daucus carota L.). J. Am. Soc. Hort. Sci. 108: 50–54.
Heineke, D., Wildenberger, K., Sonnewald, U., Willmitzer, L. and Heldt, H.W. 1994. Accumulation of hexose in leaf vacuole: studies with transgenic tobacco plants expressing yeast-derived invertase in the cytosol, vacuole or apoplasm. Planta 194: 29–33.
Klann, E.M., Chetelat, R.T. and Bennett, A.B. 1993. Expression of acid invertase gene controls sugar composition in tomato (Lycopersicon) fruit. Plant Physiol. 103: 863–870.
Laurière, C., Laurière, M., Sturm, A., Faye, L. and Chrispeels, M.J. 1988. Characterization of β-fructosidase, an extracellular glycoprotein of carrot cells. Biochimie 70: 1483–1491.
Lee, H.S. and Sturm, A. 1996. Purification and characterization of neutral and alkaline invertase from carrot. Plant Physiol. 112: 1513–1522.
Marillonnet, S. and Wessler, S.R. 1997. Retrotransposon insertion into the maize waxy gene results in tissue-specific RNA processing. Plant Cell 9: 967–978.
Miron, D. and Schaffer, A.A. 1991. Sucrose phosphate synthase, sucrose synthase, and invertase activities in developing fruit of Lycopersicon esculentum Mill. and sucrose accumulating Lycopersicon hirsutum Humb. and Bonpl. Plant Physiol. 95: 623–627.
Muller-Rober, B., Sonnewald, W. and Willmitzer, L. 1992. Inhibition of the ADP-glucose pyrophosphorylase in transgenic potatoes leads to sugar-storing tubers and influences tuber formation and expression of tuber storage protein genes. EMBO J. 11: 1229–1238.
Murray, M. and Thompson, W. 1980. Rapid isolation of highmolecular weight plant DNA. Nucl. Acids Res. 8: 4321–4325.
Simon, P.W. and Freeman, R.E. 1985. A rapid method for screening reducing sugar in carrot roots. HortScience 20: 133–134.
Simpson, C.G., Sinibaldi, R. and Brown, J.S. 1992. Rapid analysis of plant gene expression by a novel reverse transcriptase-PCR method. Plant J. 2:835–836.
Sonnewald, U., Hajirezaei, M.R., Kossman, J., Heyer, A., Tretheway, R.N. and Willmitzer, L. 1997. Increased potato tuber size resulting from apoplastic expression of a yeast invertase. Nature Biotech. 15: 794–797.
Sonnewald, U., Stitt, M., Schaewen, A.V., Brauer, M. and Willmitzer, L. 1991. Transgenic tobacco plants expressing yeastderived invertase in either the cytosol, vacuole or apoplast: a powerful tool for studying sucrose metabolism and sink/source interaction. Plant J. 1: 95–106.
Stommel, J.R. 1992. Enzymatic components of sucrose accumulation in the wild tomato species Lycopersicon peruvianum. Plant Physiol. 99: 324–328.
Stommel, J.R. and Simon, P.W. 1990. Multiple forms of invertase from Daucus carota cell cultures. Phytochemistry 29: 2087–2089.
Sturm, A. 1996. Molecular characterization and functional analysis of sucrose-cleaving enzymes in carrot (Daucus carota L.). J. Exp. Bot. 47: 1187–1192.
Sturm, A. and Chrispeels, M.J. 1990. cDNA cloning of carrot extracellular β-fructosidase and its expression in response to wounding and bacterial infection. Plant Cell 2: 1107–1119.
Sturm, A., Šebková, V., Lorenz, K., Hardegger, M., Lienhard, S. and Unger, C. 1995. Development-and organ-specific expression of the genes for sucrose synthase and three isozymes of acid β-fructofuranosidase in carrot. Planta 195: 601–610.
Tang, G.Q., Lüscher, M. and Sturm, A. 1999. Antisense repression of vacuolar and cell wall invertase in transgenic carrot alters early plant development and sucrose partitioning. Plant Cell 11: 177–189.
Unger, C., Hardegger, M., Liehard, S. and Sturm, A. 1994. cDNA cloning of carrot (Daucus carota) soluble acid β-furctofuranosidases and comparison with the cell wall isoenzyme. Plant Physiol. 104: 1351–1357.
Vivek, B.S. and Simon, P.W. 1999. Linkage relationships among molecular markers and storage root traits of carrot (Daucus carota L. ssp. sativus). Theor. Appl. Genet. 99: 58–64.
von Schaewen, A., Stitt, M., Schmidt, R., Sonnewald, U. and Willmitzer, L. 1990. Expression of a yeast-derived invertase in the cell wall of tobacco and Arabidopsis plants leads to accumulation of carbohydrate and inhibition of photosynthesis and strongly influences growth and phenotype of transgenic tobacco plants. EMBO J. 9: 3033–3044.
Wang, F., Amparo, S., Brenner, M.L. and Smith, A. 1993. Sucrose synthase, starch accumulation, and tomato fruit sink strength. Plant Physiol. 101: 321–327.
Yelle, S., Chetelat, R.T., Dorais, M., DeVerna, J.W. and Bennett, A.B. 1991. Sink metabolism in tomato fruit. IV. Genetic and biochemical analysis of sucrose accumulation. Plant Physiol. 95: 1026–1035.
Yelle, S., Hewitt, J.D., Robinson, N.L., Damon, S. and Bennett, A.B. 1988. Sink metabolism in tomato fruit. III. Analysis of carbohydrate assimilation in a wild species. Plant Physiol. 87: 737–740.
Zhang, L., Cohn, N.S. and Mitchell, J.P. 1996. Induction of a pea cell-wall invertase gene by wounding and its localized expression in phloem. Plant Physiol. 112: 1111–1117.
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Yau, YY., Simon, P.W. A 2.5-kb insert eliminates acid soluble invertase isozyme II transcript in carrot (Daucus carota L.) roots, causing high sucrose accumulation. Plant Mol Biol 53, 151–162 (2003). https://doi.org/10.1023/B:PLAN.0000009272.44958.13
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DOI: https://doi.org/10.1023/B:PLAN.0000009272.44958.13