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Zhencai Wu and Paul A. Wiersma

Expansins are a class of proteins that stimulate the extension of plant cell walls. Expansins have been found in nearly all growing plant tissues, such as hycopotyls, young seedlings, fibers, internodes, flower petals, and ripening fruits. We isolated two full-length expansin cDNA clones, Pruav-Exp1 and Pruav-Exp2, from sweet cherry (Prunus avium L.) fruit. Pruav-Exp1 has 1048 nucleotides encoding 254 amino acids, while Pruav-Exp2 has 1339 nucleotides encoding 250 amino acids. Deduced amino acid sequences of sweet cherry Pruav-Exp1 and Pruav-Exp2 share 72% identity. A Blast search of the GenBank database with the deduced amino acid sequences of Pruav-Exp1 and Pruav-Exp2 indicated a high sequence identity with other plant expansin genes. Interestingly, Pruav-Exp1 shares 99% identity of amino acid sequence with that of apricot expansin Pav-Exp1. Fragments from the 3' ends of Pruav-Exp1 and Pruav-Exp2 were cloned to generate gene-specific probes. These probes were used to study expansin gene expression in different tissues and during fruit development. Northern blot analysis showed different mRNA expression patterns for each gene. The mRNA of Pruav-Exp1 was expressed at the pink and ripe stages, but not at the early green and yellow stages of fruit development. The mRNA of Pruav-Exp2 was present earlier, from a low level in yellow expanding fruit, increasing to a high level at the pink stage and remaining at this level through the ripe stage. Both mRNAs were also expressed at a low level in flower, but not present in other tissues such as roots, leaves and peduncles. Our study indicates an expansin gene family is present in sweet cherry and suggests that two expansin genes may have different roles during fruit development and ripening.

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Paul A. Wiersma, Deniz Erogul and Shawkat Ali

Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers were evaluated in an effort to reliably DNA fingerprint sweet cherry (Prunus avium L.) cultivars and advanced selections from the breeding program at the Summerland Research and Development Center (Summerland, BC, Canada). SSR markers were found that differentiated the 35 cultivars and selections tested. However, groups of cultivars closely related to the parental cultivars, Lapins and Sweetheart, were differentiated by only a few SSR markers each. These last few markers were discovered by specifically screening within these small groups of cultivars and the resulting markers had lower discriminating power (D j) statistics within the full set of 35 cultivars and selections. To further characterize the differences in one of these closely related groups, SNP markers were identified in the cultivar Sweetheart and an analysis was made of how these markers segregated into three of its open-pollinated progeny. Large blocks of the ‘Sweetheart’ genome (34%) did not contain informative SNP markers, which was consistent with its ancestry where the cultivar Van is both a parent and grandparent. The three progeny cultivars differed from ‘Sweetheart’ at 14%, 31%, and 29% of the 3011 SNP positions tested. These were located in blocks of linked haplotypes covering from 2.5 to 20 million bps each and were distinct for the three cultivars. The cultivar Staccato®, which required the most effort for SSR marker discrimination, also had the lowest number of SNP position differences from ‘Sweetheart’ (14%). These informative SNP markers were located in only five small regions of the sweet cherry genome, which also contained the discriminating SSR markers and provides an explanation for the difficulty of locating SSR markers for this cultivar. In addition to clearly differentiating these cultivars, this SNP analysis shows the level of variation expected within this closely related group.

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Yiping Gong, Peter M.A. Toivonen, O.L. Lau and Paul A. Wiersma

Apple fruits (Malus domestica Borkh. cv. Braeburn) harvested from two orchards (A and B) on the same day were stored in air or pretreated in air for 0, 2 (2dCA) or 4 weeks (4dCA) before moving into controlled atmosphere (CA) storage with 1.5% O2 + 5% CO2. During storage at 1 °C for 9 weeks in air and/or CA, changes of pyruvate decarboxylase (PDC) activity, alcohol dehydrogenase (ADH) activity, acetaldehyde (AA) and ethanol (EtOH) concentrations in flesh tissue were assayed in addition to the incidence of Braeburn browning disorder (BBD). Immediate introduction to CA conditions induced the development of BBD with the highest incidence 62.2%, however delaying application of CA for 2 and 4 weeks reduced the incidence of BBD to 38.5% and 27.0%. The development of disorder in grower B was less than in grower A. 2dCA and 4dCA treatments did not influence PDC activity compared with treatment of CA. However, ADH activity and the accumulation of AA and EtOH in treatments of 2dCA and 4dCA were markedly lower than those in CA. The accumulation of AA in grower B was lower than grower A. The results of this study suggest that the delayed application of CA reduced BBD and this may be due to reduced anaerobic metabolism of fruits in the delayed CA.

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Peter M.A. Toivonen, Jared Stoochnoff, Kevin Usher, Changwen Lu, Paul A. Wiersma and Chunhua Zhou

The market value of the apple (Malus ×domestica Borkh.) cultivar Ambrosia is closely linked to the characteristic blush on the skin surface. For ‘Ambrosia’ orchards that produce consistently low levels of surface blush, the implementation of reflective rowcovering has improved surface coloration, but the reflected wavebands responsible for this enhanced color production have not been confirmed. This study consisted of two separate experiments: one conducted in the field to confirm reflective rowcovering efficacy and the other in a controlled environment cabinet to determine which waveband was enhancing red blush production. The red blush production in orchards with and without reflective rowcovering was then directly compared with the red blush produced on the surface of apples that were poorly colored at harvest and then exposed to visible, fluorescent, ultraviolet A (UVA), or ultraviolet B (UVB) light sources within the controlled environment chamber. Consequent analysis of the red blush color within the Commission Internationale de l’Eclairage a* and b* color space was conducted to evaluate the quality of the red blush pigment under each treatment in the field and the controlled environment chamber. The analysis revealed that the red blush that developed on apples from the reflective rowcover treatment most closely matched the red blush that developed in response to UVB exposure in the controlled environment cabinet. Further analysis of gene expression and anthocyanin contents in the ‘Ambrosia’ apples support the hypothesis that the primary driver for the characteristic red blush development, when reflective rowcovers are used, is increased exposure to UVB light.

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Lili Zhou, Frank Kappel, Cheryl Hampson, Paul A. Wiersma and Guus Bakkeren

Amplified fragment length polymorphisms (AFLPs) were used to analyze the relationships between sweet cherry (Prunus avium L.) cultivars and selections from the breeding program at the Pacific Agri-Food Research Centre in Summerland, Canada. Six pairs of preselected primers were used for the analysis of a total of 67 cultivars and selections. Scoring the absence and presence of 118 polymorphic DNA fragments produced a unique binary code for each cultivar and selection. Two phylogenetic trees were constructed using these 118 polymorphic fragments, one tree for 55 related cultivars and selections from the Summerland breeding program and the other for 23 self-incompatible cultivars of differing origins. The reliability of AFLP DNA fingerprints was confirmed by correlating relationships revealed by AFLP profiles with known genetic relationships of some sweet cherry cultivars and by a blind test for cultivar identification. Results indicate that AFLP analysis is a good technique to evaluate genetic distance and relationships in a sweet cherry breeding population.