Skip to main content
Log in

The tomato polygalacturonase gene and ripening-specific expression in transgenic plants

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

Polygalacturonase (PG) is the major cell wall degrading enzyme of tomato fruit. It is developmentally regulated and is synthesised de novo in ripening fruit. Genomic clones encoding a PG gene of tomato (Lycopersicon esculentum Mill cv. Ailsa Craig) have been isolated, mapped and sequenced. The sequence of the protein-coding region is identical to that of a PG cDNA [20]. Comparison of the cloned restriction fragments with genomic Southern data suggests that there may only be one gene for PG per haploid genome. The PG gene, which covers approximately 7 kb, is interrupted by 8 intervening sequences ranging in size from 99 bp to 953 bp. The transcription start point was identified by S1 mapping and primer extension analysis. About 1.4 kb of 5′ flanking DNA has been sequenced. This contains putative TATA and CAAT boxes and also direct repeat sequences. A transcriptional fusion has been constructed between the putative 1.4 kb promoter fragment and the chloramphenicol acetyl transferase (CAT) gene. Constructs containing this gene have been transferred to tomato using binary vectors. Regenerated transgenic plants express CAT in ripe tomato fruit, but not in unripe tomatoes, leaves, or roots.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Bernatzky R, Tanksley SD: Majority of random cDNA clones correspond to single loci in the tomato genome. Mol Gen Genet 203: 8–14 (1986).

    Google Scholar 

  2. Bevan MW: Binary Agrobacterium vectors for plant transformation. Nucleic Acids Res 12: 8711–8721 (1984).

    Google Scholar 

  3. Biggin M, Farrell PJ, Barrell BG: Transcription and DNA sequence of the Bam HI L fragment of B95–8 Epstein-Barr Virus. EMBO J 3: 1083–1090 (1984).

    Google Scholar 

  4. Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilising the principle of protein-dye binding. Anal Biochem 72: 248–254 (1976).

    Google Scholar 

  5. Breathnach R, Chambon P: Organisation and expression of eukaryotic split genes coding for proteins. Ann Rev Biochem 50: 349–383 (1987).

    Google Scholar 

  6. Brown JWS: A catalogue of splice junction and putative branch point sequences from plant introns. Nucleic Acids Res 14: 9549–9559 (1986).

    Google Scholar 

  7. Carter P, Bedouelle H, Waye MMY, Winter G: Oligonucleotide site-directed mutagenesis in M13. Anglian Biotechnology Ltd., Colchester (1985).

    Google Scholar 

  8. Chomczynski P, Qasba PK: Alkaline transfer of DNA to plastic membranes. Biochem Biophys Res Comm 122: 340–344 (1984).

    Google Scholar 

  9. Colot V, Robert LS, Kavanagh TA, Bevan MW, Thompson RD: Localisation of sequences in wheat endosperm protein genes which confer tissue-specific expression in tobacco. EMBO J 6: 3559–3564 (1987).

    Google Scholar 

  10. Dean C, Tamaki S, Dunsmuir P, Faureau M, Katayama C, Dooner H, Bedbrook J: mRNA transcripts of several plant genes are polyadenylated at multiple sites in vivo. Nucleic Acids Res 14: 2229–2240 (1986).

    Google Scholar 

  11. Della Penna D, Alexander DC, Bennett AB: Molecular cloning of tomato fruit polygalacturonase: Analysis of polygalacturonase mRNA levels during ripening. Proc Natl Acad Sci USA 83: 6420–6424 (1986).

    Google Scholar 

  12. Della Penna D, Kates DS, Bennett AB: Polygalacturonase gene expression in Rutgers, rin, nor and Nrtomato fruit. Plant Physiol 85: 502–607 (1987).

    Google Scholar 

  13. Dellaporta SL, Wood J, Hicks JB: A plant DNA minipreparation: Version 11. Plant Mol Biol Rep 1: 19–21 (1983).

    Google Scholar 

  14. Farrell PJ, Deininger P, Bankier A, Barrell BG: Homologous upstream sequences near Epstein-Barr virus promoters. Proc Natl Acad Sci USA. 80: 1565–1569 (1983).

    Google Scholar 

  15. Feinberg A, Vogelstein B: A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 137: 266–267 (1984).

    Google Scholar 

  16. Fraley RT, Rogers SG, Horsch RB, Sanders PR, Flick JS, Adams SP, Bittner ML, Brand LA, Kink CL, Fry JS, Galluppi GR, Goldberg SB, Hoffmann NL, Woo SC: Expression of bacterial genes in plant cells. Proc Natl Acad Sci USA 80: 4803–4807 (1983).

    Google Scholar 

  17. Frischauf AM, Lehrach H, Poustka A, Murray N: Lambda replacement vectors carrying polylinker sequences. J Mol Biol 170: 827–842 (1983).

    Google Scholar 

  18. Grierson D: Gene expression in ripening tomato fruit. CRC Crit Rev Plant Sci 3: 113–132 (1985).

    Google Scholar 

  19. Grierson D, Slater A, Speirs J, Tucker GA: The appearance of polygalacturonase mRNA in tomatoes: one of a series of changes in gene expression during development and ripening. Planta 163: 263–271 (1985).

    Google Scholar 

  20. Grierson D, Tucker GA, Keen J, Ray J, Bird CR, Schuch W: Sequencing and identification of a cDNA clone for tomato polygalacturonase. Nucleic Acids Res 14: 8595–8603 (1986).

    Google Scholar 

  21. Heidecker G, Messing J: Structural analysis of plant genes. Ann Rev Plant Physiol 37: 439–466 (1986).

    Google Scholar 

  22. Hobson GE: The firmness of tomato fruit in relation to polygalacturonase activity. J Hort Sci 40: 66–72 (1965).

    Google Scholar 

  23. Holdsworth MJ: Characterisation of an ethylene related small multigene family from Lycopersion esculentum. Ph.D. Thesis, University of Nottingham (1987).

  24. Holdsworth MJ, Bird CR, Ray J, Schuch W, Grierson D: Structure and expression of an ethylene related mRNA from tomato. Nucleic Acids Res 15: 731–739 (1987).

    Google Scholar 

  25. Hudson GS, Farrell PJ, Barrell BG: Two related but differentially expressed potential membrane proteins encoded by the Eco RI Dhet region of Epstein-Barr virus B95–8. J Virol 53: 528–535 (1985).

    Google Scholar 

  26. Langridge P, Feix G: A zein gene of maize is transcribed from two widely separated promoter regions. Cell 34: 1015–1022 (1983).

    Google Scholar 

  27. Maniatis T, Fritsch EF, Sambrook J: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).

    Google Scholar 

  28. Maunders MJ, Holdsworth MJ, Slater A, Knapp JE, Bird CR, Schuch W, Grierson D: Ethylene stimulates the accumulation of ripening-related mRNAs in tomatoes. Plant Cell Envir 10: 177–184 (1987).

    Google Scholar 

  29. Morelli G, Nagy F, Fraley RT, Rogers SG, Chua N-H: A short conserved sequence is involved in the light-inducibility of a gene encoding ribulose 1,5-bisphosphate carboxylase small subunit of pea. Nature 315: 200–204 (1985).

    Google Scholar 

  30. Mount SM: A catalogue of splice junction sequences. Nucleic Acids Res 10: 459–472 (1982).

    Google Scholar 

  31. Pressey R, Avants JK: Two forms of polygalacturonase in tomatoes. Biochim Biophys Acta 309: 363–369 (1973).

    Google Scholar 

  32. Proudfoot NJ, Brownlee GG: 3′ Non-coding region sequences in eukaryotic messenger RNA. Nature 263: 211–214 (1976).

    Google Scholar 

  33. Roberts JA, Schindler CB, Tucker GA: Ethylene-promoted tomato flower abscission and the possible involvement of an inhibitor. Planta 160: 159–163 (1984).

    Google Scholar 

  34. Sanger F, Nicklen S, Coulson AR: DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467 (1977).

    Google Scholar 

  35. Sheehy RE, Pearson J, Brady CJ, Hiatt WR: Molecular characterisation of tomato fruit polygalacturonase. Mol Gen Genet 208: 30–36 (1987).

    Google Scholar 

  36. Slater A, Maunders MJ, Edwards K, Schuch W, Grierson D: Isolation and characterisation of cDNA clones for tomato polygalacturonase and other ripening-related proteins. Plant Mol Biol 5: 137–147 (1985).

    Google Scholar 

  37. Sommer H, Saedler H: Structure of the chalcone synthase gene of Antirrhinum majus. Mol Gen Genet 202: 429–434 (1986).

    Google Scholar 

  38. Tucker GA, Grierson D: Synthesis of polygalacturonase during tomato fruit ripening. Planta 155: 64–67 (1982).

    Google Scholar 

  39. Tucker GA, Robertson NG, Grierson D: Changes in polygalacturonase isoenzymes during the ripening of normal and mutant tomato fruit. Eur J Biochem 112: 119–124 (1980).

    Google Scholar 

  40. Tucker GA, Robertson NG, Grierson D: The conversion of tomato-fruit polygalacturonase isoenzyme 2 into isoenzyme 1 in vitro. Eur J Biochem 115: 87–90 (1981).

    Google Scholar 

  41. Tucker GA, Schindler CB, Roberts JA: Flower abscission in mutant tomato plants. Planta 160: 164–167 (1984).

    Google Scholar 

  42. Werr W, Frommer W-B, 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 

  43. Zainon MA, Brady CJ: Purification and characterisation of the polygalacturonase of tomato fruit. Aust J Plant Physiol 9: 155–169 (1982).

    Google Scholar 

  44. Zoller MJ, Smith M: Oligonucleotide-directed mutagenesis: a simple method using two oligonucleotide primers and a single-stranded DNA template. DNA 3: 479–488 (1984).

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Bird, C.R., Smith, C.J.S., Ray, J.A. et al. The tomato polygalacturonase gene and ripening-specific expression in transgenic plants. Plant Mol Biol 11, 651–662 (1988). https://doi.org/10.1007/BF00017465

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00017465

Key words

Navigation