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Ahmad A. Omar, Wen-Yuan Song, James H. Graham and Jude W. Grosser

Citrus canker disease caused by the bacterial pathogen Xanthomonas axonopodis pv. citri is becoming a worldwide problem. Xa21 gene is a member of the Xa21 gene family of rice, which provides broad spectrum Xanthomonas resistance in rice. `Hamlin' sweet orange [Citrus sinensis (L.) Osbeck) is one of the leading commercial cultivars in Florida because of its high yield potential and early maturity. `Hamlin' also has a high regeneration capacity from protoplasts and is often used in transformation experiments. Since the citrus canker pathogen is in the same genus, this gene may have potential to function against canker in citrus. The wild-type Xa21 gene contains an intron, and there are some questions whether dicot plants can process genes containing monocot introns (the cDNA is intron-free). Plasmids DNA, encoding the non-destructive selectable marker EGFP (Enhanced Green Fluorescent Protein) gene and the cDNA of the Xa21 gene were transformed or co-transformed into `Hamlin' orange protoplasts using polyethylene glycol. More than 200 transgenic embryoids were recovered. More than 400 transgenic plants were developed from 75 independent transgenic events. PCR analysis revealed the presence of the cDNA of the Xa21 and the GFP genes in the transgenic plants. Some of the plants have the GFP only. Southern analysis is showing integration of the cDNA into different sites ranges from one to five sites. Western analysis is showing the expression of the cDNA of the Xa21 gene in the transgenic citrus plants. This is the first time that a gene from rice has been stably integrated and expressed in citrus plants. Canker challenge assay is in progress.

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Juan I. Valiente and L. Gene Albrigo

Citrus flowering is a complex phenological process influenced by a number of interacting factors. Low winter temperatures are recognized as an important factor, but the flowering response has not been quantified under Variable natural conditions. A study was conducted to monitor the flower bud induction response of `Valencia' and `Hamlin' sweet orange trees [Citrus sinensis (L.) Osbeck] to naturally occurring winter weather conditions during the 1999 and 2000 seasons. The flowering response was quantified and related to shoot age, bud position along the shoot, local weather information, and crop load status. Results indicate that buds on previous summer shoots developed 2.52 and 3.59 to 1 flower on spring shoots, for `Hamlin' and `Valencia', respectively. In addition, buds at apical positions produced more flowers than buds located far from the apex. These basal positions buds required higher induction levels. Under Florida conditions, greater accumulation of hours of temperatures 11 to 15 °C increased floral intensity by the combined effect on the number of sprouting buds with reproductive growth and the number of flowers per flowering bud. Some statistical analyses indicated that high winter temperatures reduced flowering in `Valencia' and `Hamlin' oranges. The presence of fruit consistently reduced reproductive response for both cultivars. Crop load reduced flowering by an average of 41.5% compared to no crop and varied by cultivar. A discussion on the different induction requirements as well as on the differential effect of crop load on flowering by cultivar is presented.

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Lynn Jo Pillitteri, Carol J. Lovatt and Linda L. Walling

Homologues of the floral meristem identity genes LEAFY (LFY) and APETALA1 (AP1) were isolated from the hybrid perennial tree crop `Washington' navel orange (Citrus sinensis) and designated CsLFY and CsAP1, respectively. Citrus has an extended juvenile period unlike herbaceous plants and responds to different floral stimuli than herbaceous plants or deciduous tree species. Despite these differences, the deduced amino acid sequences of CsLFY and CsAP1 genes are at least 65% identical with their Arabidopsis thaliana L. Heynh counterparts and share even greater sequence similarity to LFY and AP1 from the deciduous woody perennials, Populus balsamifera Bradshaw and Populus tremuloides Michaux, respectively. Like A. thaliana LFY (AtLFY) and AP1 (AtAP1), CsLFY and CsAP1 expression was restricted almost exclusively to reproductive tissues, but observed expression of CsAP1 in the fourth whorl carpel tissue of mature flowers was distinct from other plant AP1 genes. Transgenic A. thaliana plants ectopically expressing CsLFY or CsAP1 showed early-flowering phenotypes similar to those described for overexpression of AtLFY and AtAP1. In addition, the 35S:CsLFY and 35S:CsAP1 transgenes partially complemented the lfy-10 and ap1-3 mutants, respectively. The severity of the overexpression phenotypes correlated with the accumulation of CsLFY or CsAP1 transcripts. LFY is a single-copy gene in flowering plants but consistent with its hybrid origin, the genome of C. sinensis `Washington' has two easily distinguishable CsLFY and CsAP1 alleles derived from it's parental genotypes, C. maxima L. Osbeck (pummelo) and C. reticulata Blanco (mandarin). Allelic polymorphism at both the CsLFY and CsAP1 loci was restricted to the 5′- and 3′-flanking regions.

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Anwar G. Ali and Carol J. Lovatt

This study investigated the effect of irrigation treatments and time of foliar applications of low-biuret urea on yield of 30-yr-old navel orange trees grown under optimal N fertilization. The experiment was carried out at the Agricultural Experiment Station of the University of California, Riverside, using split plot design with 12 replications. The two irrigation treatments were the mainplots and the urea applications were the subplots. Irrigation was withheld from one set of trees from October to March, the other set was irrigated according to commercial practice. Foliar-urea (0.17 kg/tree) was applied in November, December, January, or February.

The results of the first year showed no significant differences between irrigation treatments with regard to total fruit weight or total number of fruit per tree, All trees receiving urea had significantly higher fruit weight and fruit number per tree than the control trees. The specific time of urea application had no significant effect.

With respect to fruit size distribution, the irrigated treatment resulted in a significantly higher number of fruit of size 7.0-8.0 cm (box sizes 88-72). Generally, the non-irrigatd trees had more fruit of size 6.0 cm or less. No interaction between irrigation treatments and foliar urea sprays was observed.

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Izulme R. Santos and Cecil Stushnoff

Economically, citrus is the second most-important fruit crop grown worldwide; thus germplasm conservation of commercial cultivars, as well as of wild relatives, is essential. Presently, citrus germplasm has been conserved mainly in field genebanks. This approach is helpful; however, it is costly, exposes germplasm to climatic and biological hazards, and is not a long-term conservation system. Cryopreservation (conservation in liquid nitrogen, at –150°C to –196°C) is a technique that can ensure long-term storage of plant material. Attempts to cryopreserve citrus are restricted to a few reports, but the results obtained are encouraging. The basic purpose of this study is to define cryopreservation protocols for embryo axes and axillary buds of `Pineapple' sweet orange using the encapsulation-dehydration method. Embryo axes encapsualted in Na-alginate beads, precultured with high levels of sucrose and dehydrated over silica gel before freezing in liquid nitrogen had 60% survival. No survival was obtained for buds treated the same way, however buds isolated from plants acclimated at 0°C over a 30-day period survived exposure to –20°C when slow cooled at 2°C/hour. Additional experiments will combine cold acclimation, slow cooling and pre-treatment with sugars and other chemical compounds as an attempt to enhance cold hardiness of axillary buds and obtain survival after freezing in liquid nitrogen. Different approaches will be used to increase embryo axes survival rates.

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Anwar G. Ali and Carol J. Lovatt

The ability of arginine (50 mM), putrescine (10 and 20 mM), and spermidine (10 and 20 mM) to enhance low temperature stressed-induced flowering of 5-year-old container-grown `Washington' navel orange trees was evaluated. The metabolites were applied to the foliage at the end of the 4-week low-temperature treatment of 8-h days at 10°C and 16-h nights at 7°C; the trees were then moved to warmer conditions.

All treatments had a positive effect on floral intensity. Putrescine (20 mM) and spermidine (10 mM) significantly increased (P = 0.05) total flower number and both the number and proportion of leafless inflorescences per tree. However, the number of floral shoots per tree, as well as the number of leafy inflorescences and number of vegetative shoots were not significantly influenced by the metabolites.

The results suggest that polyamines are important to the development of flowers, but not leaves, along the axis of the inflorescence.

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Craig E. Kallsen, Blake Sanden and Mary Lu Arpaia

In the southern San Joaquin Valley, early-maturing navel orange [ Citrus sinensis (L.) Osbeck] cultivars such as Beck-Earli, Newhall, and Fukumoto produce the first harvestable navel orange fruit of the season in California. Before harvest may

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Almudena Montoliu, Aurelio Gómez-Cadenas and Rosa M. Pérez-Clemente

plantings are grafted onto citranges. Materials and Methods Plant material and culture media. Three-year-old greenhouse-grown plants of the citrus rootstock Carrizo citrange [ Citrus sinensis L. Osbeck × Poncirus trifoliata L. Raf (CC)] were

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Araceli M. Vera-Guzman, Maria T. Lafuente, Emmanuel Aispuro-Hernandez, Irasema Vargas-Arispuro and Miguel A. Martinez-Tellez

.M. 2002 Abscisic acid levels and the influence of ethylene, humidity and storage temperature on the incidence of postharvest rindstaning of ‘Navelina’ orange ( Citrus sinensis (L.) Osbeck) fruit Postharvest Biol. Technol. 25 1 49 57 Lafuente, M.T. Sala, J

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Said A. Hamido, Kelly T. Morgan, Robert C. Ebel and Davie M. Kadyampakeni

schedules The experiment was conducted from 1 Jan. 2014 to 27 Aug. 2015 on 12-year-old sweet orange [ Citrus × sinensis (L.) Osbeck] trees located in three commercial groves: 1) Arcadia, FL (27.22°N, 81.86°W) in a valkaria fine sandy soil (Siliceous