Structure of the tomato Adh2 gene and Adh2 pseudogenes, and a study of Adh2 gene expression in fruit
- 124 Downloads
A cDNA library was constructed from RNA from the pericarp of ripe tomato fruit and four cDNAs encoding ADH2 were isolated and characterized. The cDNAs encode a peptide 379 amino acids in length. They hybridized strongly with a 1.8 kb RNA species well represented in RNA from ripe, but not from mature, unripe fruit, and strongly to a similar RNA species present in hypoxic, but not in aerobic roots. Northern analysis showed that the mRNA for ADH2 in fruit increased in abundance through ripening, particularly during late ripening. In pericarp tissue of fruit, the Adh2 mRNA level increased to a maximum within 8–16 h of exposure to atmospheres with 3% (v/v) oxygen, and returned to the basal level within 16 h of a return to air. The mRNA level was sensitive to the oxygen level in the atmosphere, increasing 20-fold in 12% (v/v) oxygen and 100-fold in 3% oxygen.
The homologous tomato Adh2 gene was isolated from a genomic library. The gene has an overall length of 2334 bp from transcription start site to poly(A) addition site and includes eight introns.
Southern blot analysis of tomato genomic DNA identified multiple Adh2-related sequences. Two of these, PSA1 and PSA2, were cloned and found to have 94% similarity with each other and 77% similarity with the tomato Adh2 gene over a 1000 bp region. The homologous regions include introns and exons but the equivalent exons contain frame shifts, deletions and stop codons. The two regions are therefore presumptive pseudogenes.
Key wordsADH2 cDNA fruit gene hypoxia ripening
pertaining to an ADH gene
anaerobic responsive element
Unable to display preview. Download preview PDF.
- 2.Brady CJ: Fruit ripening. Annu Rev Plant Physiol 38: 155–178 (1987).Google Scholar
- 10.Fourney RM, Miyakoshi J, Day RS, Paterson MC: Northern blotting: efficient RNA staining and transfer. Focus 10 (1): 5–9 (1988).Google Scholar
- 11.Genez AL, Staraci LC, Alexander DC, Rejda JM, Williamson VM, Chase TJnr, Williams BG: Isolation of a tomato alcohol dehydrogenase 2-encoding cDNA using phage-promoted antibody screening of a plasmid cDNA library. GenBank database, accession number M86724 (1992).Google Scholar
- 13.Gerlach WL, Pryor AJ, Dennis ES, Ferl AJ, Sacha MM, Peacock WJ: cDNA cloning and induction of the alcohol dehydrogenase gene (ADH 1) of maize. Proc Natl Acad Sci USA 79: 2981–2985 (1982).Google Scholar
- 16.Lee E, Speirs J, McGlasson WB, Brady CJ: Messenger RNA changes in tomato fruit pericarp in response to propylene, wounding or ripening. J Plant Physiol 129: 287–299 (1987).Google Scholar
- 19.Longhurst TJ, Tung HF, Brady CJ: Developmental regulation of the expression of alcohol dehydrogenase in ripening tomato fruit. J Food Biochem 14: 421–433 (1990).Google Scholar
- 20.Lyons JM, Pratt HK: Effect of stage of maturity and ethylene treatment on respiration and ripening of tomato fruits. Proc Am Soc Hort Sci 84: 491–500 (1964).Google Scholar
- 23.Messing J, Geraghty D, Heidecker G, Hnu N-T, Kridl J, Rubenstein I: Plant gene structure. In: Kosuge T, Meredith CP, Hollaender A (eds) Genetic Engineering of Plants: An Agricultural Perspective, pp. 211–227. Plenum Press, New York (1983).Google Scholar
- 24.Mozer TJ: Partial purification and characterisation of the mRNA for α-amylase from barley aleurone layers. Plant Physiol 65: 834–837 (1980).Google Scholar
- 25.Palmiter RD: Ovalbumin messenger ribonucleic acid translation. Comparable rates of polypeptide initiation and elongation on ovalbumin and globin messenger ribonucleic acid in rabbit reticulocyte lysate. Biol Chem 248: 2095–2106 (1973).Google Scholar
- 26.Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989).Google Scholar
- 28.Speirs J, Brady CJ: A coordinated decline in the synthesis of subunits of ribulosebisphosphate carboxylase in aging wheat leaves. II. Abundance of messenger RNA. Aust J Plant Physiol 8: 603–618 (1981).Google Scholar
- 31.Thomas MR, Matsumoto S, Cain P, Scott NS: Repetitive DNA of grapevine: classes present and sequences suitable for cultivar identification. Theor Appl Genet 86: 173–180 (1993).Google Scholar
- 34.Walker JC, Howard EA, Dennis ES, Peacock WJ: DNA sequences required for anaerobic expression of the maize alcohol dehydrogenase 1 gene. Proc Natl Acad Sci USA 84: 6624–6628 (1987).Google Scholar
- 35.Werr W, Frommer WB, Maas C, Starlinger P: Structure of the sucrose synthase gene on chromosome 9 of Zea mays L. EMBO J 4: 1373–1380 (1985).Google Scholar