Molecular characterization of wheat polyphenol oxidase (PPO)
- PMID: 12582641
- DOI: 10.1007/s00122-001-0847-3
Molecular characterization of wheat polyphenol oxidase (PPO)
Abstract
It is well-established that the enzyme polyphenol oxidase (PPO) is involved in undesirable browning of noodles, chapattis, middle east flat breads and steamed breads. Methods for measuring PPO activity have been developed, and the variation of PPO activity among wheat ( Triticum aestivum L.) cultivars has been well documented. However, there is no report on the identification and characterization of a wheat PPO gene. PCR performed on wheat genomic DNA with oligonucleotide primers designed from conserved copper binding regions of other PPO genes resulted in amplification of a 444-bp DNA fragment. Sequence analysis identified the conserved amino acids of PPO genes indicating that the PCR product was part of the wheat PPO gene. Screening genomic and cDNA libraries using 444- and 760-bp DNA fragments as probes failed to identify a PPO gene based on conserved sequence, even though there were very strong hybridization signals for some isolates. Rapid amplification of cDNA ends (RACE) technique was used as an alternative to obtain the remaining DNA sequences in 5' and 3' directions based on the 444-bp partial wheat PPO gene sequence. With the use of ThermoScript Reverse Transcriptase (which functions at higher temperatures) and Advantage-GC cDNA kit, the complete DNA sequence in the 3' direction was obtained. A similar effort in the 5' direction resulted in amplification of a truncated 414-bp DNA sequence. Overall, 1,509-bp of putative wheat PPO DNA sequence was obtained. Alignment of deduced amino-acid sequences revealed similarity to the other PPO gene sequences, especially in the conserved copper binding regions. Southern-blot analysis performed with four different restriction enzymes revealed two to four DNA fragments, suggesting a limited number of PPO genes in wheat. Wheat genomic DNA restricted with HindIII and hybridized using a 760-bp wheat PPO probe revealed a clear distinction between wheat cultivars with high and low PPO activities. Northern-blot analysis indicated a transcript size of about 2.0-kb. PPO DNA fragment as well as RNA transcript was observed for the durum cultivar Renville which normally has very low PPO activity. Further study is needed to explain the relationship between PPO activity and the presence of PPO gene (s).
Similar articles
-
Delineating the role of polyphenol oxidase in the darkening of alkaline wheat noodles.J Agric Food Chem. 2006 Mar 22;54(6):2378-84. doi: 10.1021/jf0526386. J Agric Food Chem. 2006. PMID: 16536622
-
Browning in Annona cherimola fruit: role of polyphenol oxidase and characterization of a coding sequence of the enzyme.J Agric Food Chem. 2007 Oct 31;55(22):9208-18. doi: 10.1021/jf070586+. Epub 2007 Oct 2. J Agric Food Chem. 2007. PMID: 17907770
-
Cloning and expression of the cathepsin F-like cysteine protease gene in Escherichia coli and its characterization.J Microbiol. 2007 Apr;45(2):158-67. J Microbiol. 2007. PMID: 17483802
-
Identification and characterisation of a bryophyte polyphenol oxidase encoding gene from Physcomitrella patens.Plant Biol (Stuttg). 2005 May;7(3):283-91. doi: 10.1055/s-2005-837598. Plant Biol (Stuttg). 2005. PMID: 15912448
-
[Analysis, identification and correction of some errors of model refseqs appeared in NCBI Human Gene Database by in silico cloning and experimental verification of novel human genes].Yi Chuan Xue Bao. 2004 May;31(5):431-43. Yi Chuan Xue Bao. 2004. PMID: 15478601 Chinese.
Cited by
-
Sequencing, assembly, annotation, and gene expression: novel insights into browning-resistant Luffa cylindrica.PeerJ. 2020 Aug 10;8:e9661. doi: 10.7717/peerj.9661. eCollection 2020. PeerJ. 2020. PMID: 32864209 Free PMC article.
-
Multifamily QTL analysis and comprehensive design of genotypes for high-quality soft wheat.PLoS One. 2020 Mar 11;15(3):e0230326. doi: 10.1371/journal.pone.0230326. eCollection 2020. PLoS One. 2020. PMID: 32160264 Free PMC article.
-
Genetic Mapping and Discovery of the Candidate Gene for Black Seed Coat Color in Watermelon (Citrullus lanatus).Front Plant Sci. 2020 Jan 22;10:1689. doi: 10.3389/fpls.2019.01689. eCollection 2019. Front Plant Sci. 2020. PMID: 32038674 Free PMC article.
-
Genome-wide linkage mapping of flour color-related traits and polyphenol oxidase activity in common wheat.Theor Appl Genet. 2016 Feb;129(2):377-94. doi: 10.1007/s00122-015-2634-6. Epub 2015 Nov 25. Theor Appl Genet. 2016. PMID: 26602234
-
Polyphenol oxidase (PPO) in wheat and wild relatives: molecular evidence for a multigene family.Theor Appl Genet. 2007 May;114(7):1239-47. doi: 10.1007/s00122-007-0514-4. Epub 2007 Feb 14. Theor Appl Genet. 2007. PMID: 17468807
LinkOut - more resources
Full Text Sources
Miscellaneous