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Bruce L. Dunn and Jon T. Lindstrom

Controlled reciprocal crosses between Trichostema lanatum Benth. (section Chromocephalum F.H. Lewis) with Trichostema arizonicum A. Gray (section Paniculatum F.H. Lewis) and Trichostema purpusii Brandegee (section Rhodanthum Lewis) were successful in generating the first artificial hybrids in the genus. Crosses where T. lanatum was used as the female were unsuccessful. Leaf and floral morphology among the hybrids was typically intermediate. Female sterility was seen in the T. arizonicum × T. lanatum hybrids, and these hybrids also produced abnormally small, nonviable pollen grains. Propagation procedures are also presented. Although these species may be in different sections, their crossability suggests that they are closely related.

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Bruce L. Dunn and Jon T. Lindstrom

Ploidy level and fertility status are often the two biggest barriers a breeder must overcome when trying to incorporate novel characteristics among related taxa. This study was aimed at developing an efficient chromosome doubling method for Buddleja L., commonly known as butterfly bush, with the goal of equalizing the ploidy level and restoring the fertility of a diploid (2n=38) F1 interspecific hybrid that has a unique orange color but happens to be sterile. This method would ease the crossing of the hybrid to the tetraploid (2n=76) B. davidii Franch. cultivars commonly found in the industry. An antimitotic treatment of oryzalin was tested on 02-25-142 (B. madagascarensis Lam. × B. crispa Benth.) in vitro using nodal sections. A factorial of varying concentrations [3, 5, and 7 μM (micromolar)] by different exposure times (1, 2, and 3 day) plus controls was set up. Oryzalin appeared to be an efficient agent for chromosome doubling in Buddleja. Significant differences in the number of polyploids were not seen between chemical concentrations and exposure times. However, higher chemical concentrations and exposure times did have a significant effect on the number of nodes that survived tissue culture. Increased leaf size and color, stem thickness, shortened internode length, and upright growth habit were all good early phenotypic indicators of polyploidy induction as later confirmed by flow cytometry. Significant increases in pollen viability accompanied chromosome doubling as crosses between 02-25-142 × B. davidii cultivars produced viable seedlings.

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Jon T. Lindstrom and Matthew C. Pelto

Itea virginica (Virginia Sweetspire) is a woody landscape shrub that has recently gained much popularity in the landscape. Several cultivars of Itea have been selected for fall leaf color and plant habit. Visual identification of some of these cultivars is difficult and confusion exists in the trade. RAPDs (randomly amplified polymorphic DNA) were used to identify Itea virginica cultivars. A single 10-base primer was sufficient to separate the cultivar Saturnalia from the cultivar Henry's Garnet. Two dwarf cultivars of I. virginica, `Merlot' and `Sprinch' (= Little Henry), were separable from `Henry's Garnet' using a single 10-base primer. A primer that distinguishes between these two dwarf cultivars has not yet been found. The technique of RAPDs appears sufficiently sensitive and repeatable to resolve questions of identity that may exist among several cultivars of Itea.

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Bruce L. Dunn and Jon T. Lindstrom

A protocol for producing fertile tetraploid forms of the hybrid Buddleja madagascarensis Lam. × B. crispa Benth. would enable introgression of orange flower, pubescence, and nondehiscent fruit characteristics found in section Nicodemia (Tenore) Leeuw. into B. davidii Franchet section Buddleja. Excised nodal sections of a single sterile diploid selection from that cross were treated in vitro with 3, 5, or 7 μm oryzalin concentrations for 1, 2, or 3 days or were left as an untreated control. A population of plants was generated from these cultures and transferred to the greenhouse. Treated plants were initially screened phenotypically for higher ploidy levels on the basis of stem thickness and leaf size. Those selected based on polyploidy characteristics were subjected to cytometric analysis, confirming that six tetraploid plants were generated. Nodal survival rates were dependent on oryzalin concentration and treatment duration. Significant increases in fertility accompanied polyploidy induction, because crosses between the newly developed tetraploids and B. davidii cultivars produced viable fertile plants. Chemical name used: 3,5-dinitro-N 4,N 4-dipropylsulfanilamide (oryzalin).

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Jon T. Lindstrom, Gregory T. Bujarski and Brent M. Burkett

Buddleja indica Lam. is encountered frequently as a houseplant or a conservatory specimen and is attractive ornamentally for its oak-shaped foliage. Buddleja indica, a tetraploid African species, 2n = 76, was crossed to the Asiatic tetraploid species B. davidii Franch. The F1 generation was intermediate in foliage character between the two parents. Flowers of the F1 were either white or light lavender in color and the number of flowers per inflorescence was intermediate between the parents. The F1 plants were fertile. These hybrids might be suitable for greenhouse or container culture due to their attractive foliage and floral display.

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Chih-Hsien Lei, Jon T. Lindstrom and William R. Woodson

At anthesis, petunia pollen contains large amounts of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). This ACC is thought to contribute to the rapid burst of ethylene produced by the pistil following pollination. An analysis of ACC content in developing anthers revealed that ACC began to accumulate the day before anthesis, indicating its synthesis was a late event in pollen development. We employed degenerate DNA primers to conserved amino acid sequences of ACC synthesis to amplify a cDNA from anther mRNA by RT-PCR. The resulting cDNA (pACS2) was sequenced and found to represent ACC synthase. Use of pACS2 as a hybridization probe revealed an increase in ACC synthase mRNA concomitant with the increase in ACC content. Further analysis indicated the ACC synthase mRNA was localized specifically to the haploid pollen grain. In an attempt to determine the function of ACC in pollen maturation or pollen–pistil interactions, we have generated a series of transgenic petunias designed to inhibit the accumulation of ACC in pollen. For these experiments, we have employed a pollen-specific promoter (LAT52) from tomato to drive the expression of antisense pACS2 or the coding region of ACC deaminase. The results of the experiments will be discussed.

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Jon T. Lindstrom, Chih-Hsien Lei and William R. Woodson

Petunia hybrida pollen accumulates significant levels of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) late in development. This pollen ACC is thought to play a role in the rapid burst of ethylene produced by pollinated pistils. To investigate this further, we have expressed the ACC deaminase gene product from Pseudomonas in transgenic petunias under the control of three different promoters including CaMV-35S, LAT52, and TA29 directing construction expression, pollen-specific expression and tapetum-specific expression, respectively. Several transgenic plants expressing the LAT52-ACC deaminase gene exhibited significant reduction of ACC in pollen. Two independent transformants contained only trace amounts of ACC in pollen. In contrast, the other promoters did not lead to reduced ACC in pollen. Pollination of wild-type pistils with pollen from LAT52-ACC deaminase plants elicited increased ethylene similar to wild-type pollen. Fecundity was unaffected by the reduction in pollen ACC content. Taken together, we conclude pollen-borne ACC is not the elicitor of pollination-induced ethylene production by pistils.

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Jon T. Lindstrom, James A. Robbins, Gerald L. Klingaman, Scott Starr and Janet Carson

The University of Arkansas established a new, replicated, woody ornamental plant evaluation program in 1999. Three sites were used across the state and these sites encompassed the three different USDA Plant Cold Hardiness Zones found in Arkansas, Zones 6, 7 and 8. In the first year, 17 different woody ornamental plants were established in the evaluation. Information obtained from performance in this evaluation will be used in Arkansas Select, a marketing program for customers and nurserymen in the state. Nonpatented and nontrademarked plant material will be made available for propagation purposes. Woody plants will be evaluated for 5 years and herbaceous perennials will be evaluated for 3 years.

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Jon T. Lindstrom, Chih-Hsien Lei, Michelle L. Jones and William R. Woodson

Mature pollen from Petunia hybrida contains significant levels of 1-aminocyclopropane-1-carboxylic acid (ACC), and this ACC is thought to play a role in pollination-induced ethylene by the pistil. We investigated the developmental accumulation of ACC in anthers and pollen. The level of ACC in anthers was very low until the day before anthesis, at which time it increased 100-fold. A 1.1-kb partial ACC synthase cDNA clone (pPHACS2) was amplified from total RNA isolated from mature anthers by reverse transcriptase, followed by polymerase chain reaction using oligonucleotide primers synthesized to conserved amino acid sequences in ACC synthases. The expression of pPHACS2 mRNA during anther development was correlated with the accumulation of ACC and was localized to the pollen grain. The pPHACS2 cDNA was used to identify the PH-ACS2 gene from a library of genomic DNA fragments from Petunia hybrida. PH-ACS2 encoded an ACC synthase transcript of four exons interrupted by three introns. The ACC synthase protein encoded by the PH-ACS2 gene shared >80% homology with ACC synthases from tomato (LE-ACS3) and potato (ST-ACS1a). A chimeric PH-ACS2 promoter-β-glucuronidase (GUS) gene was used to transform petunia and transgenic plants were analyzed for GUS activity. GUS staining was localized to mature pollen grains and was not detected in other tissues. Despite similarities to LE-ACS3, we did not detect GUS activity under conditions of anaerobic stress or in response to auxin. A series of 5-prime-flanking DNA deletions revealed that sequences within the PH-ACS2 promoter were responsible for pollen-specific expression.

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Scott E. Renfro, Brent M. Burkett, Bruce L. Dunn and Jon T. Lindstrom