Isolation and characterisation of a melon cDNA clone encoding phytoene synthase

Abstract

A cDNA clone (MEL5), encoding a protein homologous to phytoene synthase (PSY), has been isolated from a climacteric melon fruit cDNA library, using the tomato cDNA clone TOM5 [34] as a heterologous probe. MEL5 hybridised to a transcript of 1.65 kb which suggested that the 1.36 kb clone, isolated originally, was not full-length. The missing 5' end was isolated by a reverse transcriptase-polymerase chain reaction (RT-PCR)-based method. This enabled the full sequence of the protein to be deduced and the cleavage site of the transit peptide for chromoplast import to be predicted. Northern analysis of RNA extracted from fruit samples of different ripening stages as well as from roots, leaves and flower petals was used to examine the expression pattern of the corresponding mRNA. The corresponding mRNA. The transcript corresponding to MEL5 is present at low quantities in unripe (green) fruit, reaches its highest levels when the fruit turns from green to orange and persists at lower levels during later ripening stages. A similar transcript was also detected in flower petals and in trace amounts in leaves and roots. Genomic Southern analysis indicates that the clone is homologous to a low-copy-number gene family. Sequence analysis showed a high degree of conservation among plant PSYs.

A comprehensive review on the colorless carotenoids phytoene and phytofluene

“…In rice, poplar, tomato, wheat and maize there are two or more homolog genes, whereas only one has been identified in Arabidopsis [51][52][53][54][55][56][57][58][59][60][61][62][63]. The presence of two or more PSY enzymes seems to be important in the control of carotenoid biosynthesis in different organs, at different developmental stages or in response to environmental stimuli, including stresses like salt and drought [27].…”

CHRC, Encoding a Chromoplast-specific Carotenoid-associated Protein, Is an Early Gibberellic Acid-responsive Gene

“…A random priming kit (Boehringer Mannheim) was used to radioactively label DNA probes. The blots were hybridized with 32 P-labeled cucumber CHRC cDNA (16) and reprobed with melon phytoene synthase (PSY) cDNA (MEL5) (18), kindly provided by Prof. D. Grierson (Nottingham University, UK). The hybridization for analysis of CHRC expression was carried out in 0.263 M Na 2 HPO 4 , 7% SDS, 1 mM EDTA, 1% BSA for 16 h at 60°C, and the washes were performed in 2 ϫ SSC/0.1% SDS at 50°C followed by 2 ϫ SSC/0.1% SDS at 55°C, for 20 min each.…”

Color genes in the orchid Oncidium Gower Ramsey: identification, expression, and potential genetic instability in an interspecific cross

“…In a number of other examples of carotenoid-accumulating flowers, the expression of the carotenoid genes, including PSY, paralleled the formation of carotenoids. Examples include the yellow flowers of tomato, melon, and Gentiana lutea (Giuliano et al 1993;Karvouni et al 1995;Zhu Northern membrane was exposed for 14 days et al 2002). In daffodil flowers, the PSY gene was upregulated relative to leaves and remained elevated and constant while the PSY protein accumulated (Schledz et al 1996).…”