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P. Gercheva, R.H. Zimmerman, L.D. Owens, C. Berry, and F.A. Hammerschlag

Shoot regeneration from apple (Malus domestica Borkh.) leaf explants following particle bombardment at various acceleration pressures was studied. Basal leaf segments of micropropagated `Royal Gala' apple were bombarded with 1 μm gold particles, accelerated using helium pressures of 4.5, 6.2, 7.6, 9.3, or 13.8 MPa (650–2000 psi), and cultured on shoot regeneration medium consisting of N6 salts supplemented with 10 μM TDZ for 5, 10, or 20 days in darkness. Bombarded and control explants exhibited 63% to 100% shoot regeneration. With a 5-day dark period, average shoot production per explant ranged from 6.1 to 14; bombardments of 4.5 and 6.2 MPa significantly increased shoot production over the controls. With a 10-day dark period, average shoot production per explant ranged from 9.1 to 22 following bombardment at 9.3 and 6.2 MPa, respectively. Following bombardment at 6.2 MPa, 75% of the explants produced more than 20 regenerants per explant. With a 20-day dark period, average shoot production per explant ranged from 8.9 to 19 following bombardment at 13.8 MPa and following no bombardment, respectively. Shoot production per explant was significantly less than the controls following bombardments ranging from 6.2 to 13.8 MPa. Shoot production was highest per explant with particle bombardment at 6.2 MPa followed by incubation in darkness for 10 days. Chemical name used: thidiazuron (TDZ).

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D.S. Tustin, T. Fulton, and H. Brown

Growth of apple fruit can be described as an initial exponential phase lasting the 40+ days of fruit cell division followed by a more-or-less linear phase where growth is by cell expansion. Temperature is a major influence on fruit growth rate during the cell division phase, thereby affecting fruit size at maturity. However it is generally thought that temperature has less-direct impact on fruit development during the fruit expansion phase. Our observations of apple growth among regions and seasons of considerable climatic variability led us to speculate that temperature may impact directly on fruit development during fruit expansion but that responses may be interactive with carbon balance (crop load) influences. Controlled environment studies are being used to examine this hypothesis. Potted `Royal Gala' trees set to three levels of crop (one fruit per 250, 500, or 1000 cm2 leaf area) were grown from 56 to 112 DAFB in day/night temperature regimes of 18/6, 24/12, and 30/18 °C. All trees grew in field conditions prior to and following the controlled environment treatments. Treatments were harvested when 20% to 25% of fruit on trees showed the visual indicators used commercially to indicate harvest maturity. Fruit were evaluated using attributes that determine quality and that may have implications for fruit post harvest behaviour. Temperature and crop load influences on time to maturity, fruit fresh and dry weight, fruit DM content, fruit firmness, fruit airspace content and estimated fruit cortical cell size will be presented and implications discussed.

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I.J. Warrington, T.A. Fulton, E.A. Halligan, P.T. Austin, A.J. Hall, and P.W. Gandar

Apple fruitlet growth responses to temperature were studied, for different durations following bloom (DAFB), under controlled environment (CE) conditions. Container-grown trees of `Red Delicious', `Golden Delicious', `Braeburn', `Fuji', and `Royal Gala' were placed in different maximum/minimum temperature regimes, ranging from 9/3 to 25/15°C for various periods, including 10–40, 10–80, and 40–80 DAFB. Temperature treatments were selected to identify possible differences between mean and maximum/minimum differential effects Trees were placed outdoors following the CE treatment to allow impacts on subsequent fruit development to be determined. The impact of temperature was dramatic. For example, fruit expansion rate for `Red Delicious' varied from 0.12 mm/day at 9/3°C to 0.98 mm/day at 25/15°C. Furthermore, the cell division phase was considerably longer under cooler temperatures. The influence of post-bloom temperature, for even short durations, was evident at harvest in both fruit size and in different fruit maturity indices. Differences in temperature sensitivity were evident amongscultivars. A detailed model has been developed to integrate the responses that have been determined.

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Jayasree Ravi, Donn T. Johnson, Barbara A. Lewis, and Curt R. Rom

In 1995, greenhouse and orchard experiments of 11 apple cultivars were conducted in Fayetteville and Clarksville, Ark. Weekly cumulative mite days (CMD) were regressed against leaf bronzing (colorimeter value L) and compared among cultivars. European red mites, Panonychus ulmi and two-spotted spider mites, Tetranychus urticae, were found on leaves. `Liberty', `Royal Gala', and `Stark Spur Red Rome Beauty' had significantly more mites (>1940 CMD) than did `Arkansas Black' (1303), `Jonafree' (1150), and `Northern Spy' (973). A low CMD on `Northern Spy' caused leaves to bronze faster [y = 29.04 + 0.006(x); R 2 = 52, P = 0.0002] than did a high CMD on `Liberty' [y = 30.41 + 0.0027(x); R 2 = 70, P = 0.0001]. Field estimates were made of spider mites/leaf and bronzing from 20 June to 7 Aug. `Stark Spur Red Rome Beauty' and `Stark Spur Law Rome' had significantly more CMD than did `Northern Spy' and `Arkansas Black'. Apple cultivars differed in carrying capacity to mites (susceptibility) and how fast leaves bronze in response to mite feeding. Cultivar differences in hairiness of the lower leaf surface were not correlated to CMD.

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Jorge B. Retamales, Claudio Valdes, and Verónica Donoso

Bitter pit (BP) is the main physiological disorder of apples in Chile. Its incidence affects pre- and postharvest handling of the fruit and the profitability of this species. Since 1991, its control and prediction have been studied by this research team through field and laboratory trials. The BP incidence is linked to the fruit Ca concentration; however, fruit Ca analysis has not adequately predicted BP incidence in postharvest. Several authors have proposed Ca/Mg antagonism, which has been the basis to develop a predictive method through fruit Mg infiltration (IMG) 40 days before harvest. IMG has been massively used for two seasons in Chile, with 375 samples processed in 1997 and 1170 in 1998. The industry has been increasing its proportion of the samples processed, from 22% in 1997 to 71% in 1998. The most prominent varieties are `Granny Smith' (GS) > `Braeburn' (BR) > `Royal Gala' (RG) > `Red King Oregon' (RKO). The massive use of IMG has obtained predictive capacities (r 2 between BP predicted and BP after 3 months regular cold storage) of 0.8 for `Fuji'; 0.7 for GS, BR, and RG; and 0.58 for RKO. (This reduction in the predictive capacity with regards to the previous research under controlled conditions would, in part, be due to problems in obtaining fruit samples: non-uniform fruit size, inadequate sampling dates, diverse fruit numbers, etc.) Developments are underway to increase the geographical coverage of the service, the predictive capacity of the method for certain cultivars and productive areas and the number of samples processed.

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Rudaina Alrefai, Schuyler S. Korban, and Leslie L. Domier

Two strains of Agrobacterium tumefaciens carrying a disarmed Ti-binary vector, pZA-7, were used as vectors for transformation of apple leaf segments, EHA101:pZA-7 carries a helper plasmid derived from pTiB0542, and C58Z707:pZA-7 carries a helper plasmid derived from pTiC58. The binary vector provides two selection markers, neomycin phosphotransferase (nptII) and hygromycin resistance genes, and a screening marker, β-glucuronidase (GUS) gene. Preliminary experiments were conducted to determine the effect of different concentrations of kanamycin, carbenicillin, and cefotaxime on regeneration of apple leaf sections and inhibition of A. tumefaciens strains. In vitro-derived leaf sections of `Royal Gala' apple were grown on a regeneration medium containing thidiazuron and NAA; these were then dipped into a suspension culture of A. tumefaciens and transferred to a fresh regeneration medium. Callus lines exhibiting kanamycin and hygromycin resistance were obtained mostly with agrobacterium strain EHA101:pZA-7. Expression of GUS activity was also determined in putative transformed calli. Southern blot analysis was used for confirming integrative transformation in transgenic lines.

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I.J. Warrington, T.A. Fulton, E.A. Halligan, and H.N. de Silva

Container-grown `Delicious', `Golden Delicious', `Braeburn', `Fuji' and `Royal Gala' apple [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.] trees, on Malling 9 (M.9) rootstock, were subjected to a range of different maximum/minimum air temperature regimes for up to 80 days after full bloom (DAFB) in controlled environments to investigate the effects of temperature on fruit expansion, final fruit weight, and fruit maturation. Fruit expansion rates were highly responsive to temperature with those at a mean of 20 °C being ≈10 times greater than those at a mean of 6 °C. All cultivars exhibited the same general response although `Braeburn' consistently showed higher expansion rates at all temperatures compared with lowest rates for `Golden Delicious' and intermediate rates for both `Delicious' and `Fuji'. The duration of cell division, assessed indirectly by measuring expansion rate, appeared to be inversely related to mean temperature (i.e., prolonged under cooler conditions). Subsequently, fruit on trees from the coolest controlled temperature treatment showed greater expansion rates when transferred to the field and smaller differences in fruit size at harvest than would have been expected from the measured expansion rates under the cool treatment. Nonetheless, mean fruit weight from warm postbloom treatments was up to four times greater at harvest maturity than that from cool temperature treatments. Postbloom temperature also markedly affected fruit maturation. Fruit from warm postbloom temperature conditions had a higher soluble solids concentration, more yellow background color, lower flesh firmness, and greater starch hydrolysis than fruit from cooler temperatures.

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Preston K. Andrews, David J. Chalmers, and Mapasaka Moremong

Temperature differences between tree canopies and air (Tc - Ta) and between leaves and air (T1 - Ta) of apples (Malus domestics Borkh. `Royal Gala') grown in New Zealand were measured with infrared (IR) thermometry. Treatments included three orchard-floor management systems and irrigation withheld (WI) for part of the growing season. Measurements of soil moisture indicated that, under full irrigation (FI), an alfalfa orchard-floor system apparently had higher soil water content than herbicide-strip (H) or plastic-mulch systems, whereas under the drought stress of WI, the H system retained the most water. The Tc - Ta and T1 - Ta of the WI treatment were significantly greater than those of the FI treatment after a soil-moisture differential was established. Linear regression between Tc - Ta, or T1 - Ta, and vapor pressure deficit (VPD) exhibited variable responses among dates. A crop water stress index (CWSI) was calculated from environmental measurements. The calculated CWSIS were not related to soil-moisture measurements. Even 35 days after full irrigation had been reinstated on the WI plots, the Tc - Ta, T1 - Ta, and CWSI of the WI plots were still significantly greater than those of the FI plots. These discrepancies in IR thermometry-based water-stress indices may be due to increased errors in the calculation of minimum CWSI at low VPDS and to fluctuating solar radiation and evapotranspiration, which are prevalent in humid, temperate climates.

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Freddi A. Hammerschlag, Richard H. Zimmerman, Umedi L. Yadava, Sally Hunsucker, and Petya Gercheva

A range of antibiotics and short-term exposure to an acidified (pH 3.0) medium were evaluated for their effects on eliminating Agrobacterium tumefaciens, supervirulent strain EHA101 (pEHA101/pGT100), from leaf explants of `Royal Gala' apple (Malus ×domestica Borkh.) and on shoot regeneration. Exposure of leaf explants to regeneration and elongation media containing 100 μg·mL-1 concentrations of the antibiotics carbenicillin (crb), cefotaxime (cef), and cefoxitin [=mefoxin (mef)], singly or in combination for 52 days did not eliminate A. tumefaciens from the explants. The percentage of regeneration on crb, cef, and mef was 97%, 11%, and 50%, respectively, compared to 67% for the controls. Short-term (1- to 18-hour) vacuum infiltration with 500 μg·mL-1 of any of the above antibiotics did not inhibit regeneration and failed to eliminate A. tumefaciens from leaf explants. Cef (2000 μg·mL-1) did not inhibit the percentage of regeneration and was more effective than crb or mef in preventing growth of A. tumefaciens when vacuum infiltrated into apple leaf explants for 30 minutes. Further experiments demonstrated that the incidence of A. tumefaciens contamination could be reduced to 28% without negatively impacting shoot regeneration by using a 1-hour vacuum infiltration with an acidified medium, an 18-hour vacuum infiltration with cef (5000 μg·mL-1), and a 52-day incubation on regeneration and elongation media containing 100 μg·mL-1 each of mef and crb. Kan resistant, GUS (β-glucuronidase) positive, putative transformants without A. tumefaciens were generated by adding kan (10 μg·mL-1) to the regeneration and elongation media.

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Yi Tan, Baisha Li, Yi Wang, Ting Wu, Zhenhai Han, and Xinzhong Zhang

species, leaves of 28 cultivars or stocks had regeneration rates that ranged between 5% and 100% ( Sun et al., 2000 ). An apple genotype, GL-3, was screened out from 100 in vitro seedling clones of Royal Gala with both 100% regeneration capacity and the