Molecular analysis of natural populations of Populus nigra L intermingled with cultivated hybrids

In this study six simple sequence repeats (SSR or microsatellites) were selected for their ability to fingerprint a total of 60 commercial clones of Populus deltoides Marsh. and Populus x canadensis Moench (typically derived from crosses between Populus nigra L and P. deltoides) and to characterize a natural population of P. nigra growing along the Ticino river in the North of Italy. Out of six SSRs used, four microsatellite loci were found to have alleles which were species-specific to P. deltoides and could therefore be used as markers for introgression of P. deltoides into P. nigra. In the studied region hybrid poplars and P. deltoides commercial clones are cultivated as monoclonal stands close to the area where black poplar has its natural habitat. SSR analysis was performed to investigate whether there was evidence of introgression between the natural population and the monoclonal plantations of hybrids and P. deltoides clones cultivated in the surrounding area. Three stages of the natural population were analysed: a group of old trees about a hundred years old, a younger population (aged 2-30 years) and the seedlings of three females of this population. Alleles specific to P. deltoides were detected only in the old cohort of the natural population, while no introgression was observed in the younger individuals and their progenies. These results were also confirmed by isozyme analysis of loci PGI-B, PGM and LAP-A, which were previously identified as diagnostic for P. nigra, P. deltoides and P.xcanadensis.     

Factors affecting Agrobacterium tumefaciens-mediated transformation in several black poplar clones

Transient expression of the uidA reporter gene was used in preliminary experiments with two oncogenic and two disarmed Agrobacterium tumefaciens strains in order to test the efficiency of T-DNA transfer to N084 x Populus nigra and N107 x P. nigra clones. The oncogenic strain A281 pKIWI105 produced the highest average number of GUS spots per leaf disc. In order to optimize the production of transgenic plantlets from different P. nigra clones (San Giorgio, Jean Pourtet, N084 x P. nigra and N107 x P. nigra, respectively), two A. tumefaciens strains (GV2260 p35S GUS, A281pKIWI105) and bacterial concentrations (7×10⁸; 1.2×0⁹ bacteria ml^-1) were used. Following co-cultivation with A281 pKIWI105, the frequency of leaf discs producing kanamycin-resistant calli was not significantly different between the clones and bacteria concentrations used. Transformed shoots were regenerated from all clones, except for Jean Pourtet. Co-cultivation of leaf discs with GV2260 p35S GUS produced very few calli which died when transferred to selective regeneration medium. In addition, the effects of acetosyringone and leaf wounding were evaluated for the San Giorgio and Jean Pourtet clones, using the same strains. Factors which significantly affected the transformation efficiency of leaf explants were the P. nigra clone, the A. tumefaciens strain, and the presence of acetosyringone. Genetic transformation of calli and regenerated plantlets was confirmed by their ability to grow and root on Woody Plant Medium containing kanamycin, by histochemical -glucuronidase assays, and Southern blot hybridization analyses.Abbreviations BA benzyladenine - GUS -glucuronidase - IBA indolebutyric acid - MS Murashige and Skoog - NAA -naphthaleneacetic acid - nptII neomycin phosphotransferase II gene - uidA -glucuronidase gene - WPM Woody Plant Medium     

Evaluating the tolerance of young hybrid poplar trees to recycled waters high in salinity and boron

The successful adoption of water recycling strategies in many arid regions will require crops able to tolerate poor-quality waters. We evaluated different clones for salt and boron (B) tolerance within each of seven genetically distinct genomic groups (e.g., deltoides, deltoides x nigra, trichocarpa x deltoides, trichocarpa x deltoides x maximowizcii, trichocarpa x deltoides x nigra, trichocarpa x nigra, trichocarpa x maximowizcii). During each evaluation period, different clones within each of the groups were irrigated with high sodium chloride (NaCl) salinity (i.e., 10-30 dS m(-1)) and B (i.e., 10 mg L(-1)) water up to a maximum of 150 days, for a 4-year testing period under micro-field plot conditions. Excessive accumulation (up to 6%) of chloride (Cl) likely caused toxicity symptoms (necrosis of the leaves) observed in the less tolerant clones, while leaf B concentrations rarely exceeded 300 mg kg(-1) DM in any clone. Increased soil salinity likely hindered the uptake of B by the clones. Our results show that a wide range of selected Populus clones, of parentage trichocarpa x nigra, followed by deltoides x nigra show potential salt and B tolerance as young trees to recycled waters high in salinity and B.     

Molecular genetic analysis of black poplar (Populus nigra L.) along Dutch rivers

The genetic structure of remaining black poplar ( Populus nigra ) trees on the banks of the Dutch Rhine branches was investigated using the AFLP technique. In total, 143 trees, including one P. deltoides and some P. x euramericana , were analysed using six AFLP primer combinations which generated 319 polymorphic bands. The AFLP patterns showed that some of the trees sampled as P. nigra were clearly different. These deviating patterns were also observed for the P. deltoides tree and all trees already identified as hybrid P. x euramericana . Hybrids between the two species are morphologically sometimes difficult to distinguish from the species itself. Two important possible source populations for recolonization of the riverbanks of the river Rhine, consisting of mature flowering P. nigra trees, appeared to consist of only a few genotypes each. In contrast, young black poplar trees growing alone or in small groups downstream of the possible source populations appeared to be predominantly generatively derived because no clones of mature trees were found among them. Therefore vegetative propagation seems a very local strategy whereas colonization of new areas appears to occur through generative propagation. Whether the genetic diversity within these black poplars is sufficient for recolonization of river banks and survival of the metapopulation is a question for further research.     

Infiltration with Agrobacterium tumefaciens induces host defense and development-dependent responses in the infiltrated zone

Despite the widespread use of Agrobacterium tumefaciens to transfer genes into plant systems, host responses to this plant pathogen are not well understood. The present study shows that disarmed strains of Agrobacterium induce distinct host responses when infiltrated into leaves of Nicotiana tabacum. The responses are limited to the infiltrated zone and consist of i) induction of pathogenesis-related (PR) gene PR-1 expression and resistance to subsequent infection with tobacco mosaic virus, ii) chlorosis and loss of chloroplast rRNAs, and iii) inhibition of leaf expansion. Induction of the latter two sets of responses depends on the age of the leaf and is most apparent in young leaves. Strains with or without binary vectors induce all the responses, showing that DNA transfer is neither required nor inhibitory. A. tumefaciens cured of the tumor-inducing (Ti) plasmid is slightly defective for induction of the three responses, showing that Ti plasmid-encoded factors produced by the disarmed strains contribute only slightly. However, T-DNA-encoded factors alter at least one of the host responses, because infiltration with the oncogenic strain C58 induced more pronounced chlorosis than the disarmed control. Auxin is one of the T-DNA products responsible for disease induction by oncogenic A. tumefaciens. We found that C58-infiltrated zones-but not those infiltrated with the disarmed control-have increased levels of miR393, a microRNA that represses auxin signaling and contributes to antibacterial resistance.     

Matrix attachment regions (MARs) enhance transformation frequency and transgene expression in poplar

We tested the value of a matrix attachment region (MAR) fragment derived from a tobacco gene for increasing the frequency of Agrobacterium-mediated transformation. A binary vector that carried a GUS reporter gene containing an intron and an nptII gene was modified to contain flanking MAR elements within the T-DNA borders. Vectors containing or lacking MARs were then used to transform tobacco, a readily transformabl e poplar clone (Populus tremula × P. alba), and a recalcitrant poplar clone (Populus trichocarpa × P. deltoides). MARs increased GUS gene expression approximately 10-fold in the two hybrid poplar clones and twofold in tobacco one month after cocultivation with Agrobacterium; MARs also increased the frequency of kanamycin-resistant poplar shoots recovered     

Influence of poplar clones on fertility life-table parameters of Chaitophorus leucomelas (Hemiptera: Aphididae)

The aphid Chaitophorus leucomelas Koch (Hemiptera: Aphididae) is one of the most important pests of poplar (Populus spp.) plantations in Iran. In this study, development, reproduction, and life history of the aphid were assessed on 11 poplar clones; belong to three species, Populus nigra L., Populus deltoides Bartram ex Marshall, and Populus. euramericana Guinier. The experiments were carried out under controlled conditions at 24 +/- 1 degrees C, 50-60% RH, and a photoperiod of 12:12 (L:D) h. The developmental time at immature stage ranged from 10 to 12 d. Nymphs reproduced per female were ranged from 49 to 98 nymphs on Populus deltoides var. missoriensis and P. deltoides 72/51, respectively. The intrinsic rate of natural increase (r(m)) varied from 0.176 to 0.264 d(-1) among poplar clones. The r(m) values of the aphids were adversely affected in P. euramericana 242 in comparison with P. nigra 56/72 and P. nigra 63/135. In addition, the jackknife estimates of net reproductive rate (R0) indicated the presence of resistance among poplar clones. R0 ranged from 16.48 on P. nigra var. betulifoli to 47.53 on P. nigra 63/135. Mean generation times (T) was last 17.56 d on P. euramericana 242 to 14.51 d on P. deltoides 69/55. However, doubling time (DT) was 3.87 d on P. euramericana var. grandis to 2.63 d on P. nigra 63/135. The finite rate of increase (lambda) was 1.192 on resistant clone (P. euramericana 242) and 1.302 on susceptible clone (P. nigra 63/135). These results indicate that variation in life-table parameters could be an important component of variation in resistance to C. leucomelas in poplar.     

Enhanced soybean infection by the legume "super-virulent" Agrobacterium tumefaciens strain KAT23

Agrobacterium tumefaciens KAT23 harbors a nopaline-type Ti plasmid and is "super-virulent" to soybean (Glycine max) and other leguminous plants. The right and left border sequences of the essential cis-element for T-DNA transfer were removed in order to utilize the high infectivity of this strain in an Agrobacterium-mediated soybean transformation system. The resulting strain, named Soy2, showed no oncogenic activity. After inoculation with disarmed Soy2 harboring binary vector pIG121-Hm and pCAMBIA-WR, soybean epicotyls exhibited high beta-glucuronidase activities, with efficiencies higher than EHA105, an A. tumefaciens strain widely used in making transgenic plants.     

Genetic transformation of Populus nigra by Agrobacterium tumefaciens

Two clones of Populus nigra L. were tested in vivo and in vitro for their susceptibility to three different Agrobacterium tumefaciens wild-type strains evaluating number and size of resulting calluses. Strain C58 proved to be the most virulent.Various parameters affecting Agrobacterium-mediated transformation of P. nigra clones were further analyzed using ß-glucuronidase gene transient expression. The clone Jean Pourtet proved to be more susceptible than the clone San Giorgio. A. tumefaciens strain A281 pKIWI105 proved to be the most virulent. The optimal procedure involved dipping of leaf discs into a bacterial suspension (7×10⁸ cells/ml) for 20 min, followed by a 48 h co-cultivation period on semi-solid regeneration medium.Leaf explants were co-cultivated with two disarmed A. tumefaciens strains. Plantlets of San Giorgio were regenerated, tested for ß-glucuronidase activity and rooted on selective medium containing kanamycin. Polymerase chain reaction analysis and Southern blot hybridization confirmed the integration of the neomycin phosphotransferase II gene into the poplar genome.Abbreviations BAP 6-benzyl-aminopurine - CaMV Cauliflower Mosaic Virus - 2,4-D 2,4-dichlorophenoxyacetic acid - GUS and gus ß-glucuronidase - hpt hygromycin phosphotransferase - IBA indole-3-butyric acid - KIN kinetin - LB Luria Bertani - MS Murashige and Skoog - NAA ßnaphthaleneacetic acid - NOS Nopaline synthase - NPTII and nptII neomycin phosphotransferase II - PCR Polymerase chain reaction - PVC poly-vinyl-cloride - SDS sodium dodecyl sulfate - SSC sodium cloride-sodium citrate - Tris tris(hydroxymethyl)amino-methane - WPM Woody Plant Medium     

Screening hybrid poplar clones for susceptibility to Cryptorhynchus lapathi (Coleoptera: Curculionidae)

The poplar-and-willow borer, Cryptorhynchus lapathi (L.) (Coleoptera: Curculionidae), is a wood-boring pest of economic importance in irrigated hybrid poplar (Populus spp.) farms in eastern Washington and Oregon. There is no practical insecticide control tactic against either the larval or adult stage of C. lapathi. To assess variability in C. lapathi toward clone preference, we initiated a no-choice study on 180 caged trees that consisted of five clones in a randomized complete block design. C. lapathi was significantly more successful at establishing a population in two clones with Populus trichocarpa X P. deltoides (TxD) parentage (female x male) than in either of two clones with P. deltoides x P. nigra (DxN) parentage (female x male), or a single clone with P. deltoides x P. maximowiczii (DxM) parentage (female X male). There were no significant differences in the rate of weevil development among infested clones, with the exception of DxM trees. Larvae in DxM clones developed on average to the fourth size grouping and those in the two TxD clones developed on average to the fifth size grouping, and this difference was significant. These results corroborate our general damage surveys conducted in the field. Our findings provide growers with the option to choose less susceptible varieties when replanting.     

Hybrid poplar clones with Populus maximowiczii parentage demonstrate postoviposition antibiosis to Cryptorhynchus lapathi (Coleoptera: Curculionidae)

Damage caused by the poplar-and-willow borer, Cryptorhynchus lapathi (L.) (Coleoptera: Curculionidae), is reported to vary among hybrid poplar clones. We evaluated oviposition preferences and larval success in four hybrid poplars on potted and field-planted trees. Oviposition occurred somewhat less frequently and abundantly on two clones with Populus maximowiczii Henry parentage in field-planted and potted trees, and significantly fewer larvae survived to adulthood on those clones. No adults emerged from field-planted NM 6 (Populus nigra L. x P. maximowiczii) and four emerged from TM 256-28 (Populus trichocarpa Torrey & Gray x P. maximowiczii) on which male-female pairs of C. lapathi had been caged. In contrast, 50 and 140 adults emerged over the same 2-yr period from two susceptible clones (n = 20), TD 52-226 (P. trichocarpa x Populus deltoides Bartram ex Marshall) and TN 302-9 (P. trichocarpa x P. nigra), respectively. Thus, resistance expressed by clones with P. maximowiczii parentage partially involves decreased levels of oviposition, but more significantly, antibiosis in resistant clones prevents the development of larvae, probably in early spring.     

美洲黑杨不同无性系对分月扇舟蛾幼虫的抗性及相关机理

为了选育出在生产上具有推广价值的美洲黑杨Populus deltoides优良抗虫品系,本研究以分月扇舟蛾Clostera anastomosis 3龄幼虫为材料,以I-72杨为对照评价和分析了18个美洲黑杨无性系在实验室条件（28±1℃,RH 70%,16L∶8D）下的抗虫性,通过选择性和非选择性取食实验筛选出了11个具有较强抗性的无性系; 测定了美洲黑杨主要营养物质和次生代谢物质含量,研究了不同无性系对分月扇舟蛾体内保护酶和消化酶活性及对该虫生长发育和食物利用的影响。结果表明：美洲黑杨不同无性系抗性的大小与它们叶片中总酚含量呈明显正相关趋势; 抗性水平不同的无性系对分月扇舟蛾幼虫的生长发育和食物利用有不同的影响,表现在幼虫体重、每日体重增加量、相对生长率、近似消化率、食物利用率和食物转化率等主要生长发育指标的变化上,同时对分月扇舟蛾幼虫取食量和排粪量亦有不同的影响。结果说明,分月扇舟蛾对次生代谢物质的反应更为敏感,抗性无性系对分月扇舟蛾幼虫体内SOD和CAT酶活性有明显的激活作用。     

Mitochondrial DNA variation and genetic relationships of Populus species.

We examined variation in and around the region coding for the cytochrome c oxidase I (coxI) and ATPase 6 (atp6) genes in the mitochondrial genomes of four Populus species (P. nigra, P. deltoides, P. maximowiczii, and P. tremuloides) and the natural hybrid P. x canadensis (P. deltoides x P. nigra). Total cellular DNAs of these poplars were digested with 16 restriction endonucleases and probed with maize mtDNA-specific probes (CoxI and Atp6). The only variant observed for Atp6 was interspecific, with P. maximowiczii separated from the other species as revealed by EcoRI digestions. No intraspecific mtDNA variation was observed among individuals of P. nigra, P. maximowiczii, P. x canadensis, or P. tremuloides for the CoxI probe. However, two varieties of P. deltoides were distinct because of a single site change in the KpnI digestions, demonstrating that P. deltoides var. deltoides (eastern cottonwood) and var. occidentalis (plains cottonwood) have distinct mitochondrial genomes in the region of the coxI gene. Populus x canadensis shared the same restriction fragment patterns as its suspected maternal parent P. deltoides. Nucleotide substitutions per base in and around the coxI and atp6 genes among the Populus species and the hybrid ranged from 0.0017 to 0.0077. The interspecific estimates of nucleotide substitution per base suggested that P. tremuloides was furthest removed from P. deltoides and P. x canadensis and least diverged from P. nigra. Populus maximowiczii was placed between these two clusters.     

Agrobacterium mediated transformation and regeneration of Populus

Summary A plant transformation and regeneration system has been developed for Populus species. Leaf explants, from stabilized shoot cultures of a Populus hybrid NC-5339 (Populus alba x grandidentata), were co-cultivated with Agrobacterium tumefaciens on a tobacco nurse culture. Both oncogenic and disarmed strains of A. tumefaciens harboring a binary vector which contained two neomycin phophotransferase II (NPT II) and one bacterial 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase (aroA) chimeric gene fusions were used. Shoots did not develop when leaf explants were co-cultivated with the binary disarmed strain of A. tumefaciens. However, transformed plants with and without the wild type T-DNA were obtained using an oncogenic binary strain of A. tumefaciens. Successful genetic transformation was confirmed by NPT II enzyme activity assays, Southern blot analysis and immunological detection of bacterial EPSP synthase by Western blotting. This is the first report of a successful recovery of transformed plants of a forest tree and also the first record of insertion and expression of a foreign gene of agronomic importance into a woody plant species.     

Removal of nitrogen and phosphorus by Eucalyptus and Populus at a tertiary treated municipal wastewater sprayfield

Various progenies of Eucalyptus grandis and E. amplifolia, and clones of Populus deltoides, were evaluated for plant removal of nitrogen (N) and phosphorus (P) for 26 months at a municipal waste spray field in north Florida. Tertiary treated wastewater containing 2.73 mg L(-1) nitrate N and 0.30 mg L(-1) total P was applied using sprinkler irrigation (93.8 m3 ha(-1) d(-1)) to fast growing trees utilized for bioenergy. Eucalyptus amplifolia and E. grandis survived and grew very poorly as the result of severe winter injury in two successive years and were not evaluated for nutrient removal. Survival and growth of P. deltoides demonstrated suitability for phytoremediation, and selected clones were evaluated for biomass and nutrient content. Removals of total N (TN) and total P (TP) were greatest for main stem (36% and 44%, respectively) and foliage (44% and 36%, respectively). Low biomass producing clones generally had higher nutrient concentrations, but high biomass producing clones removed more TN and TP. Approximately 789 kg ha(-1) TN and 103 kg ha(-1) TP were removed by the highest biomass producing P. deltoides clone, representing 215% of N and 615% of P inputs.     

Repeated unidirectional introgression towards Populus balsamifera in contact zones of exotic and native poplars

As the evolutionary significance of hybridization is largely dictated by its extent beyond the first generation, we broadly surveyed patterns of introgression across a sympatric zone of two native poplars ( Populus balsamifera , Populus deltoides ) in Quebec, Canada within which European exotic Populus nigra and its hybrids have been extensively planted since the 1800s. Single nucleotide polymorphisms (SNPs) that appeared fixed within each species were characterized by DNA-sequencing pools of pure individuals. Thirty-five of these diagnostic SNPs were employed in a high-throughput assay that genotyped 635 trees of different age classes, sampled from 15 sites with various degrees of anthropogenic disturbance. The degree of admixture within sampled trees was then assessed through Bayesian clustering of genotypes. Hybrids were present in seven of the populations, with 2.4% of all sampled trees showing spontaneous admixture. Sites with hybrids were significantly more disturbed than pure stands, while hybrids comprised both immature juveniles and trees of reproductive age. All three possible F1s were detected. Advanced-generation hybrids were consistently biased towards P. balsamifera regardless of whether hybridization had occurred with P. deltoides or P. nigra. Gene exchange between P. deltoides and P. nigra was not detected beyond the F1 generation; however, detection of a trihybrid demonstrates that even this apparent reproductive isolation does not necessarily result in an evolutionary dead end. Collectively, results demonstrate the natural fertility of hybrid poplars and suggest that introduced genes could potentially affect the genetic integrity of native trees, similar to that arising from introgression between natives.     

Dense genetic linkage maps of three Populus species (Populus deltoides, P. nigra and P. trichocarpa) based on AFLP and microsatellite markers

Populus deltoides, P. nigra, and P. trichocarpa are the most important species for poplar breeding programs worldwide. In addition, Populus has become a model for fundamental research on trees. Linkage maps were constructed for these three species by analyzing progeny of two controlled crosses sharing the same female parent, Populus deltoides cv. S9-2 x P. nigra cv. Ghoy and P. deltoides cv. S9-2 x P. trichocarpa cv. V24. The two-way pseudotestcross mapping strategy was used to construct the maps. Amplified fragment length polymorphism (AFLP) markers that segregated 1:1 were used to form the four parental maps. Microsatellites and sequence-tagged sites were used to align homoeologous groups between the maps and to merge linkage groups within the individual maps. Linkage analysis and alignment of the homoeologous groups resulted in 566 markers distributed over 19 groups for P. deltoides covering 86% of the genome, 339 markers distributed over 19 groups for P. trichocarpa covering 73%, and 369 markers distributed over 28 groups for P. nigra covering 61%. Several tests for randomness showed that the AFLP markers were randomly distributed over the genome.     

An improved procedure for production of white spruce (Picea glauca) transgenic plants using Agrobacterium tumefaciens.

An efficient and reproducible procedure for the transformation of white spruce (Picea glauca [Moench] Voss) embryogenic tissues was developed using A. tumefaciens-mediated gene transfer. Rapidly dividing white spruce embryogenic tissues were co-cultivated with disarmed A. tumefaciens strains containing additional copies of the virulence regions from plasmid PToK47. The plasmid pBi121, containing the neomycin phosphotransferase II (nptII) gene providing kanamycin resistance as a selectable marker and the beta-glucuronidase (uidA) reporter gene, was used as binary vector. The highest frequency of transformation (15 transformed tissues g(-1) FW of treated embryogenic tissue) was obtained with 5-d-old tissues grown in liquid medium and co-cultivated with Agrobacterium for 2 d in the same medium but containing 50 microM acetosyringone. Recovery of kanamycin-resistant tissues was improved when tissues were first grown for 10 d on a timentin-containing medium (400 mg l(-1)), to prevent bacterial overgrowth, before application of the selection pressure. After 6 weeks on kanamycin-selection medium, resistant tissues were obtained and showed stable uidA expression. The presence of the transgenes was demonstrated by PCR analysis and their integration into the genome was confirmed by Southern hybridization. Transgenic plants were regenerated from transformed tissues within 4 months after co-culture.     

Differential susceptibility of poplar hybrids to the aphid Chaitophorus leucomelas (Homoptera: Aphididae)

After its recent introduction to Chile, the aphid Chaitophorus leucomelas Koch is becoming a serious pest affecting commercial poplar, Populus spp., plantations. The pattern of natural infestation of C. leucomelas among poplar hybrids with different pedigrees and the aphid intrinsic rate of increase (r(m)), of C. leucomelas were assessed in the field. In most of the hybrids, aphid abundance peaked in March (late summer). Among 12 types of poplar crosses, [(P. trichocarpa Torr. & Gray x P. deltoides Bartram ex Marshall) x (P. trichocarpa x P. deltoides)] and [(P. trichocarpa x P. maximowiczii Henry) x P. maximowiczii] showed the highest and lowest aphid densities, respectively. A trend to find more aphids in branch bases was apparent. The intrinsic rate of C. leucomelas increase was higher in [(P. trichocarpa x P. deltoides) x P. deltoides] hybrids, and lower in [(P. trichocarpa x P. maximowiczii) x P. trichocarpa] hybrids. Aphid density and performance were higher in hybrids with P. deltoides parentage, whereas hybrids with P. maximowiczii parentage showed lower aphid densities and performance. Hybrids with P. nigra L. parentage, namely, [P. trichocarpa x P. nigra], also had high aphid density, but aphid performance was lower compared with hybrids with P. deltoides parentage. These results suggest that among poplar hybrids studied, susceptibility to C. leucomelas is inherited through P. deltoides, whereas resistance seems to be inherited through P. maximowiczii. Thus, P. maximowiczii hybrids are recommended for commercial or ornamental planting programs in zones where there is a high risk of aphid infestation.     

Agrobacterium-mediated transformation of African violet

A method for genetic transformation of Saintpaulia ionantha by co-cultivation of in vitro-grown leaves and petioles with Agrobacterium tumefaciens is described. Two bacterial strains, EHA105 and A281 both harbouring the binary plasmid pKIWI105 carrying the genes uidA and nptII, were used in the experiments. Regenerants were not obtained using the disarmed strain EHA105. The oncogenic strain A281 resulted in efficient transient and stable expression of the transferred traits for petiole explants only. After transformation and regeneration, the integration of the transgenes in the plant genome was confirmed by PCR analysis and Southern hybridization.