Expression of the murine ornithine decarboxylase gene in transgenic Nicotiana tabacum var xanthi

Ornithine decarboxylase (ODC), arginine decarboxylase (ADC), and S-adenosyl-methionine decarboxylase (SAMDC) are the three key regulatory enzymes for polyamine (putrescine, spermidine, and spermine) biosynthesis. In order to gain more insight into the relationship between polyamine metabolism and other physiological processes, research was undertaken to obtain increased putrescine biosynthesis in tobacco by overexpression of a murine ODC cDNA. Both a full-length and a truncated murine ODC cDNA were cloned into a binary expression vector containing the Cauliflower Mosaic Virus (CaMV) 35S promoter. Using standard leaf-disc transformation procedures, transgenic tobacco plants containing either the full length or the truncated ODC cDNA were obtained. Presence of the murine ODC cDNA as well as transcription were confirmed via Southern and Northern blotting. Western blot analysis identified a polypeptide unique to the transformed plants which immunoreacted with anti-ODC antibody. A series of enzyme assays were done to differentiate between native and murine ODC activity. Assays were run at the pH optima for native ODC (pH 8.2) and murine ODC (pH 6.8). At pH 6.8, there was very little activity in the control plants, but a significantly higher activity in the transformed plants. Difluoromethylornithine (DFMO), a specific irreversible inhibitor of ODC activity, completely inhibited ODC activity in the transformed plants at pH 6.8. However in the control plants at pH 6.8 and both the control and transformed plants at pH 8.2, DFMO only inhibited ODC activity by approximately 30-50%. Almost 100% inhibition of ODC activity by immunoprecipitation of ODC protein with anti-ODC antibody was observed in the transformed plants, at pH 6.8. The results show clearly that the activity of murine ODC can be detected and quantified even in the presence of the plant ODC. The transgenic plants containing the truncated ODC cDNA always had several fold higher activity than those containing the full-length cDNA. Transgenic plants containing the truncated ODC cDNA contained 10-12 times the levels of putrescine than the control plants. Transgenic plants containing the full-length cDNA contained 4-5 times the level of putrescine as compared to the control. In addition to increased levels of putrescine, there was an amine-containing compound unique to the transformed plants with a retention time very similar to putrescine

Characterization of Novel Di-, Tri-, and Tetranucleotide Microsatellite Primers Suitable for Genotyping Various Plant Pathogenic Fungi with Special Emphasis on Fusaria and Mycospherella graminicola

The goals of this investigation were to identify and evaluate the use of polymorphic microsatellite marker (PMM) analysis for molecular typing of seventeen plant pathogenic fungi. Primers for di-, tri-, and tetranucleotide loci were designed directly from the recently published genomic sequence of Mycospherlla graminicola and Fusarium graminearum. A total of 20 new microsatellite primers as easy-to-score markers were developed. Microsatellite primer PCR (MP-PCR) yielded highly reproducible and complex genomic fingerprints, with several bands ranging in size from 200 to 3000 bp. Of the 20 primers tested, only (TAGG)4, (TCC)5 and (CA)7T produced a high number of polymorphic bands from either F. graminearum or F. culmorum. (ATG)5 led to successful amplifications in M. graminicola isolates collected from Germany. Percentage of polymorphic bands among Fusarium species ranged from 9 to 100%. Cluster analysis of banding patterns of the isolates corresponded well to the established species delineations based on morphology and other methods of phylogenetic analysis. The current research demonstrates that the newly designed microsatellite primers are reliable, sensitive and technically simple tools for assaying genetic variability in plant pathogenic fungi

The Garage Band Lightshow

The expenses musicians accrue make it hard to afford all the different equipment needed to perform and very unlikely that the average start up band or artist can afford the luxury of a musical light show in their early performances. The Garage Band Lightshow provides an easily set up and affordable way for musicians to use concert lighting in their performances. The Garage Band Lightshow provides one LED box. The light box responds to an audio signal and provides varying pulses and colors of light depending on the high, mid, and low frequencies of the audio signal. The Garage Band Lightshow provides a professional concert lightshow without the use of expensive software or a light technician. Instead, the musician can create the custom lightshow by simply playing music

Modulation of the polyamine biosynthetic pathway in transgenic rice confers tolerance to drought stress

We have generated transgenic rice plants expressing the Datura stramonium adc gene and investigated their response to drought stress. We monitored the steady-state mRNA levels of genes involved in polyamine biosynthesis (Datura adc, rice adc, and rice samdc) and polyamine levels. Wild-type plants responded to the onset of drought stress by increasing endogenous putrescine levels, but this was insufficient to trigger the conversion of putrescine into spermidine and spermine (the agents that are believed to protect plants under stress). In contrast, transgenic plants expressing Datura adc produced much higher levels of putrescine under stress, promoting spermidine and spermine synthesis and ultimately protecting the plants from drought. We demonstrate clearly that the manipulation of polyamine biosynthesis in plants can produce drought-tolerant germplasm, and we propose a model consistent with the role of polyamines in the protection of plants against abiotic stress

Intersporal Genetic Variation of Gigaspora margarita, a Vesicular Arbuscular Mycorrhizal Fungus, Revealed by M13 Minisatellite-Primed PCR

Spores of vesicular arbuscular mycorrhizal (VAM) fungi contain thousands of nuclei. In order to understand the karyotic structure of a VAM fungus spore, the genetic variation of the first generation of spores from a VAM fungus (Gigaspora margarita) was examined. Spores originating from both single- and multispore inoculations of the species G. margarita were analyzed by M13 minisatellite-primed PCR. In both cases, different fingerprints were obtained from individual spores with few spores exhibiting similar fingerprints. These results can be explained only by a heterokaryotic status of the nuclear population within a spore