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Mark Ehlenfeldt and Ronald L. Prior

Antioxidant capacity as measured by ORAC, total phenolic, and total anthocyanin concentrations were evaluated in leaf tissue of the same 86 highbush blueberry cultivars, and ORAC and phenolic levels evaluated in leaf tissue of the same materials. Average values for ORAC, phenolics, and anthocyanins in fruit were 15.9 ORAC units (1 unit = 1 μmol Trolox Equivalent), 1.79 mg/g (gallic acid equivalents), and 0.95 mg/g (cyanidin-3-glucoside equivalents), respectively. `Rubel' had the highest ORAC values, at 31.1 units. Values for ORAC and phenolics in leaf tissue were significantly higher than fruit tissue, with mean values of 490.4 ORAC units and 44.8 mg/g in leaf tissue, respectively. No significant correlations were found between fruit ORAC and leaf ORAC, or between fruit ORAC and leaf phenolics. Investigation of ORAC values in a family of 44 `Rubel' × `Duke' seedlings showed negative epistatis for ORAC values. However, an analysis of ORAC values vs. pedigree in plants from the 86 cultivar groups suggested that, across cultivars, ORAC inheritance in generally additive.

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I.L. Goldman

Wisconsin Fast Plants are rapid-cycling versions of various Brassica species amenable to a variety of genetic studies due to their short life cycle and ease of handling. I have recently developed a model system using Brassica rapa L. Fast Plants for teaching the cyclical selection process known as recurrent selection in the context of a course on plant breeding. The system allows for up to three cycles of recurrent selection during a 15-week semester and enables students to gain experience in planting, selection, pollination, and seed harvest during each cycle. Fourteen cycles of replicated, recurrent mass selection for high (H) and low (L) levels of anthocyanin pigment expression in hypocotyl tissue were practiced by students in Horticulture 502 during a period of four semesters. In addition to bi-directional selection; replicated unselected (D) control populations were maintained forcomparative purposes. Over 14 cycles, highly significant gains and losses in hypocotyl pigment production were realized for H and L populations, respectively. Plants in D populations showed no directional response to random selection and therefore did not exhibit genetic drift. Plants in H populations exhibited production of anthocyanin pigment in organs other than hypocotyls, suggesting selection goals could be modified to include pigmentation of specific organs or whole plants. Results from this selection program suggest significant gains from recurrent selection can be visualized through student-based selection activities in the classroom.

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Mark K. Ehlenfeldt and Ronald L. Prior

Antioxidant capacity as measured by ORAC, total phenolic, and total anthocyanin concentrations were evaluated in fruit tissue of 86 highbush blueberry cultivars, and ORAC and phenolic levels evaluated in leaf tissue of the same materials. Average values for ORAC, phenolics, and anthocyanins in fruit were 15.9 ORAC units (1 unit = 1 micromole Trolox Equivalent), 1.79 mg·g–1 (gallic acid equivalents), and 0.95 mg·g–1 (cyanidin-3-glucoside equivalents), respectively. ëRubel' had the highest ORAC values, at 31.1 units. Values for ORAC and phenolics in leaf tissue were significantly higher than fruit tissue, with mean values of 490.4 ORAC units and 44.8 mg·g–1 in leaf tissue, respectively. No significant correlations were found between fruit ORAC and leaf ORAC, or between fruit ORAC and leaf phenolics. Investigation of ORAC values in a family of 44 `Rubel' × `Duke' seedlings showed negative epistasis for ORAC values. However, an analysis of ORAC values vs. pedigree in plants from the 86 cultivar groups suggested that across cultivars, ORAC inheritance is generally additive.

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Sunggil Kim*, Marla Binzel, Sunghun Park, Kil-Sun Yoo and Leonard Pike

Anthocyanin, one of the flavonoids, is a primary determinant of red color in onions. Inheritance studies indicate that a single gene determines the color difference between yellow and red onions. In order to establish which gene might be responsible for this color difference, full-length cDNAs of five structural genes: chalcone synthase (CHS), flavanone 3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and flavonol synthase (FLS) were cloned using degenerate PCR and RACE (Rapid Amplification of cDNA Ends). RT-PCR was carried out for these five genes to examine differential expression between yellow and red colored bulbs. Accumulation of the DFR gene transcript only occurred in red onions. In F3 populations which originated from the cross between yellow and red parents, DFR transcript was detected only in red F3 lines, not in yellow F3 lines. To design molecular markers for selection of yellow and red DFR alleles, the DFR gene was sequenced from genomic DNA isolated from both types of onions. The genomic DNA sequence revealed the DFR gene consists of six exons and five introns. A PCR-RFLP marker was designed based on 2% polymorphic nucleotide sequence of the DFR gene between yellow and red onions. The co-segregation of markers and red color were observed in F2 segregating populations, supporting the conclusion that color difference in red and yellow onions is likely to be due to the lack of an active DFR gene.

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Yi hu Dong, Deepali Mitra, Arend Kootstra, Carolyn Lister and Jane Lancaster

The red color of Royal Gala apple (Malus domestics Borkh.) skin increased in intensity following irradiation with ultraviolet (UV) and white light. The enhanced red apple color was due to an increase in anthocyanin concentration and the increase was dose dependent. High-performance liquid chromatography analysis showed that the composition of flavonoids in UV treated and natural red colored apple skins was similar. The red apple skin color further increased after storage at 4C in the dark. During the course of irradiation the enzymatic activities of phenylalanine ammonia lyase (PAL) and chalcone isomerase (CHI) increased 10-to 20-fold. Northern analysis showed an increase in PAL transcripts during the irradiation treatment, suggesting that the increase in PAL enzymatic activity was due to de novo synthesis of the enzyme in apple skin cells.

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W.J. Steyn, D.M. Holcroft, S.J.E. Wand and G. Jacobs

Changes in activity of phenylalanine ammonia-lyase (PAL) and UDPGalactose: flavonoid-3-o-glycosyltransferase (UFGT) during the development of pear (Pyrus communis L.) fruit and in response to cold fronts were assessed and related to changes in red color. Red and blushed pear cultivars attained maximum redness and highest anthocyanin concentrations in immature fruit. Red color generally faded toward harvest. UFGT activity increased over fruit development and was apparently not limiting to color development. However, the fading of red color and the decreasing level of phenolic compounds toward harvest might relate to decreasing PAL activity. Skin color and enzyme activity in the red pear `Bon Rouge' displayed little responsiveness to low temperatures. In contrast, low temperatures increased red color and activity of both PAL and UFGT in the blushed pear `Rosemarie'. Consistent with the general pigmentation pattern described above, the effect of temperature on enzyme activity was much greater early during fruit development than in the week before harvest.

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Ann Marie Connor, Chad E. Finn, Tony K. McGhie and Peter A. Alspach

Dietary anthocyanins (ACYs) may give health benefits through their antioxidant activity (AA) or other physiological effects. We examined AA and ACY profiles and contents in 16 blackberry and hybridberry (Rubus L. species) cultivars harvested in 2002 and 2003 in New Zealand and Oregon. Total ACY content varied widely among cultivars harvested from a single site in a single year (e.g., from 58 to 343 mg/100 g fruit Oregon in 2003). For the 12 cultivars common to both sites and years, cultivar and year within location significantly affected total ACY content, accounting for 40% and 10% of total variation, respectively. Cultivar interactions with both location and year within location were also significant, together accounting for 39% of variation. Cyanidin 3-O-glucoside and cyanidin 3-O-rutinoside were identified in all cultivars in both locations in at least 1 year. Compared with total ACY, cultivar accounted for more variation in these two ACYs (63% and 92%, respectively), while cultivar interactions together accounted for a smaller, but statistically significant, proportion of variation (23% and 7%, respectively). Cyanidin 3-O-sophoroside and cyanidin 3-O-(2G-glucosylrutinoside) were identified in only four cultivars. Cultivar effects accounted for 64% and 76% of variation in these ACYs, respectively, while cultivar interactions together contributed 18% and 24%, respectively. For AA, cultivar effects were not significant, contributing 11% of variation; in contrast, year effect and cultivar × environment interactions were significant, contributing 22% and 55% of total variation, respectively. Based on cultivar means for all 16 genotypes, the phenotypic correlation between AA and total ACY was positive but lower than that between AA and total phenolic content (TPH) (r = 0.63 and 0.97, respectively). Combinations of individual ACYs did not provide higher correlations with AA. Thus, ACY profiles and content are not as useful as TPH as a proxy measurement for AA.

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Ann Marie Connor, Tony K. McGhie, M. Joseph Stephens, Harvey K. Hall and Peter A. Alspach

We determined variance components and narrow-sense heritability estimates for total and individual anthocyanin (ACY) content and antioxidant activity (AA) in fruit from 411 genotypes in a red raspberry (Rubus idaeus L.) factorial mating design based on 42 full-sib families derived from seven female and six male parents, harvested in 2002 and 2003. Within half-sib family total ACY content ranged from ≈1-60+ mg/100 g fruit in both seasons. The four major ACYs quantified by high-performance liquid chromatography also showed wide ranges each year. Female and male parent contributions to variation in total and individual ACYs were significant (P ≤ 0.001) in combined year analysis, and together accounted for 29% to 48% of the total variation. A substantial proportion of the female contribution was attributed to the use of a pigment-deficient R. parvifolius L. × R. idaeus hybrid derivative as a female parent. Female × male interaction was nonsignificant and contributed negligibly to total variance. Year effects accounted for <2.5% of variation in ACYs and were only marginally significant. Year interactions were negligible. Within family variation (among plots and within plot) accounted for ≈50% of the variation in total ACY and 62% to 69% of the variation in individual ACYs. Combined year narrow-sense heritability estimates were high (h 2 = 0.54-0.90 for individual ACYs, 1.00 for total ACY) among all factorial genotypes, but moderate when the progeny of the R. parvifolius derivative were excluded (h 2 = 0.45-0.78 for individual ACYs, 0.74 for total ACY). The latter estimates are applicable to breeding programs in which pigment-deficient genotypes are rarely or never used in breeding. Parental main effects were significant for AA, together accounting for 19% of total variance; female × male interaction was nonsignificant. Year effects were marginally significant and year interactions nonsignificant; together these sources of variation contributed <2% of total variation in AA. The majority of AA variation was found within- and among-plots within family. The phenotypic correlation between AA and total ACY was r = 0.53, and ranged from r = 0.21-0.46 between AA and individual ACYs; genetic correlations between AA and the ACYs were similar to the phenotypic correlations, suggesting predominantly additive genetic effects accounted for the phenotypic correlations. Linear modelling for AA based on individual ACYs and their interactions explained ≈0.53 of AA variation, substantially less than that explained by total phenolic content (R 2 = 0.88). Our results show substantial variation and moderate to high narrow-sense heritability estimates for red raspberry ACYs, but ACY content and profile information are ineffective proxies and predictors for AA in red raspberry fruit.

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Milton E. Tignor, Frederick S. Davies, Wayne B. Sherman and John M. Davis

Poncirus trifoliata (L.) Raf. seeds were germinated in perlite under intermittent mist at about 25 °C and natural daylight in a greenhouse. Two-week-old seedlings were then transferred into a growth chamber at 25 °C and 16-hour daylength for 1 week. Tissue samples were collected at 0, 6, 24, 168, and 504 hours after temperature equilibration at 10 °C. Freezing tolerance at –6.7 °C, as determined by electrolyte leakage, and stem (leaves attached) water potential (ψx), measured using a pressure chamber, was recorded for a subset of seedlings for each time interval. Red coloration (apparently anthocyanin) developed at the petiole leaflet junction and buds after 48 hours at 10 °C and gradually occurred throughout the leaves during further exposure. Complementary DNA clones for phenylalanine ammonia lyase (PAL), 4-coumarate: coA ligase (4CL), and chalcone synthase (CHS) were used to probe RNA isolated from the leaves. No increase in steady-state messenger RNA level was detected. Increases in freeze hardiness occurred within 6 hours in the leaves, and continued for up to 1 week. Water potential initially decreased from –0.6 to –2.0 MPa after 6 hours, then returned to –0.6 MPa after 1 week. Thus, Poncirus trifoliata seedlings freeze-acclimate significantly after only 6 hours at 10 °C.

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D.L. Deal, J.C. Raulston and L.E. Hinesley

Red- and purple-leafed seedlings and clonal material selected for superior color and growth under northern climatic conditions may exhibit progressive color loss and reduced growth rates when exposed to the hot summers and high night temperatures of more southern climates. Studies were conducted to characterize the color loss associated with red-leafed seedlings of Acer palmatum Thunb. (Japanese maple), and to determine to what extent night temperatures affect the dark respiration, growth, and anthocyanin expression of A. palmatum `Bloodgood'. The percentage of seedlings within each of five color classes was determined for five dates from spring to early fall. Significant shifts in class distribution occurred on every evaluation date tested. The class changes contributing the most to these shifts varied with age of leaf material and date. Dark respiration rates increased by 0.09 mg CO2/g leaf dry weight per hour for every 1C rise in temperature, regardless of exposure duration. Dark respiration rates of 0.69 and 1.73 mg CO2/g per hour were found at 14 and 26C, respectively. The greatest amount of growth occurred during weeks 6 through 8 at a night temperature of 14C. Plant growth during this period increased by an average 51%, compared to that at warmer night temperatures. Ultimately, total plant growth at 14C decreased 7%, 19%, and 32% as night temperatures increased from 18 to 22 to 26C. Leaf redness index values at 14 or 18C were from two to seven times greater than those at warmer night temperatures.