Summary
The completion of the Arabidopsis thaliana genome has revealed that there are nine members of the Pht1 family of phosphate transporters in this species. As a step towards identifying the role ...of this gene family in phosphorus nutrition, we have isolated the promoter regions from each of these genes, and fused them to the reporter genes β‐glucuronidase and/or green fluorescent protein. These chimeric genes have been introduced into A. thaliana, and reporter gene expression has been assayed in plants grown in soil containing high and low concentrations of inorganic phosphate (Pi). Four of these promoters were found to direct reporter gene expression in the root epidermis, and were induced under conditions of phosphate deprivation in a manner similar to previously characterised Pht1 genes. Other members of this family, however, showed expression in a range of shoot tissues and in pollen grains, which was confirmed by RT‐PCR. We also provide evidence that the root epidermally expressed genes are expressed most strongly in trichoblasts, the primary sites for uptake of Pi. These results suggest that this gene family plays a wider role in phosphate uptake and remobilisation throughout the plant than was previously believed.
Phosphorus (P) enters roots as inorganic phosphate (Pi) derived from organic and inorganic P compounds in the soil. Nucleic acids can support plant growth as the sole source of P in axenic culture ...but are thought to be converted into Pi by plant-derived nucleases and phosphatases prior to uptake. Here, we show that a nuclease-resistant analog of DNA is taken up by plant cells. Fluorescently labeled S-DNA of 25 bp, which is protected against enzymatic breakdown by its phosphorothioate backbone, was taken up and detected in root cells including root hairs and pollen tubes. These results indicate that current views of plant P acquisition may have to be revised to include uptake of DNA into cells. We further show that addition of DNA to Pi-containing growth medium enhanced the growth of lateral roots and root hairs even though plants were P replete and had similar biomass as plants supplied with Pi only. Exogenously supplied DNA increased length growth of pollen tubes, which were studied because they have similar elongated and polarized growth as root hairs. Our results indicate that DNA is not only taken up and used as a P source by plants, but ironically and independent of Pi supply, DNA also induces morphological changes in roots similar to those observed with P limitation. This study provides, to our knowledge, first evidence that exogenous DNA could act nonspecifically as signaling molecules for root development.
Citrus black spot caused by
is characterized by fruit blemishes and premature fruit drop, resulting in significant economic losses in summer rainfall areas. The pathogen forms both conidia and ...ascospores during its life cycle. However, the occurrence of these spores and their contributions to infection of fruit in field conditions are not well-understood. Our research using direct leaf litter monitoring and volumetric spore trapping in Queensland orchards revealed that pseudothecia and ascospores in leaf litter, as well as trapped ascospores had low abundance, while pycnidia and conidia were highly abundant. Both
and endophytic
spp. were identified, with
being dominant. In replicated field trials we determined that infection of 'Imperial' mandarin fruit by
occurred from fruit set until week 20 of fruit development, with the key infection events taking place between weeks 4 and 16 in Queensland subtropical conditions. These results demonstrate that protecting fruit during weeks 4 to 16 significantly reduced
infection. We found no significant correlation between the disease incidence in fruit and
conidial abundance in leaf litter, or ascospore abundance measured by volumetric spore trapping. It is therefore suggested that inoculum sources in the tree canopy other than those detected by spore trapping and direct leaf litter monitoring may play a major role in the epidemiology of citrus black spot. The improved knowledge on epidemiology of
and an understanding of propagules causing infection may aid in development of more effective disease management strategies.
Key message
The AGPase large subunit (shrunken-2) promoter was demonstrated to be active in the placentochalaza and endosperm of developing grain as well as the root tips in transgenic sorghum.
The ...temporal and spatial expression patterns of the
Sorghum bicolor Shrunken
-
2
(
Sh2
) promoter were evaluated using the green fluorescence protein reporter gene (
gfp
) in transgenic sorghum, within the context of upregulating starch biosynthesis in the developing grain. GFP fluorescence was analysed throughout development in various tissue types using confocal laser scanning microscopy techniques.
Sh2
promoter activity was first detected in the placentochalaza region of the developing caryopsis and apoplasm adjacent to the nucellar epidermis at 7 days post anthesis (dpa) where fluorescence remained relatively constant until 17 dpa. Fluorescence in this region weakened by 20 dpa and disappeared by 25 dpa. Expression was also detected in the developing endosperm, but not until 12 dpa, continuing until 25 dpa. Whilst the endosperm expression was expected, the fluorescence detected in the placentochalaza was completely unexpected. Although transcript presence does not mean the resulting biochemistry is also present, these preliminary findings may suggest alternate spatial activity of ADP-glucose pyrophosphorylase prior to uptake by the developing grain.
Sh2
promoter activity was also unexpectedly detected in the root tips at all developmental time points.
Sh2
promoter activity was not detected in any reproductive floral tissue (both pre and post anthesis) or in pollen. Similarly, no expression was detected in leaf tissue at any stage.
An unknown sclerotium-forming fungus was observed and isolated in 2016 from sugarcane grown at Sugar Research Australia, Woodford Pathology farm, Woodford, Queensland. The fungus appeared to be ...causing germination failure of sugarcane setts. The fungus was isolated from infected tissues and mycelia from internal pithy sett tissues. White mycelia and abundant sclerotia were grown on PDA. Morphological observation and molecular analysis confirmed the identification of the fungus as
Athelia rolfsii
. Significant reductions in sett germination (>70%) were observed in the field when inoculated setts were planted. Reddish and patchy lesions were observed on the external surface of the setts and pale-red to red discolouration in internal tissues (associated with white mycelial growth in the pithy tissues and on the cut end of setts). Symptoms on young plants included water-soaked to light-brown lesions at their base, production of sclerotia / white mycelium in and around the lesions, and death of the young plants. This is the first report demonstrating that
A. rolfsii can
cause germination failure and young plant death in sugarcane.
The efficient use of transgenic technology for the improvement of phosphorus (P) nutrition in crop plants may require the expression of introduced genes only under conditions of phosphate deficiency ...and only in specific tissues. We have investigated the use of the promoter from the
Arabidopsis
Pht1;2 phosphate transporter to drive the expression of a secretable
Aspergillus niger phytase gene only in the root epidermis of phosphate-deprived plants. Deletion analysis of this promoter indicated that, while the 933 bp upstream of the start codon is sufficient for root-specific and phosphate-responsive reporter gene expression, the 2000 bp upstream of the start codon are required for maximum expression levels. We show that transgenic
Arabidopsis plants in which this 2000 bp promoter drives the
Aspergillus phytase gene secrete phytase enzyme only from roots when grown on medium containing low phosphate concentrations, and that this transgene enables the plants to grow on medium containing phytate as a sole P source. The growth rates and shoot P concentrations of these plants were similar when grown on phytate or phosphate as the P source, and were similar to transgenic lines in which the phytase was driven by the constitutive CaMV 35S promoter. Our results demonstrate the potential usefulness of the promoter from the
Pht1;2 phosphate transporter for driving genes that may improve crop P uptake and nutrition.
Phosphate transport in plants Smith, Frank W.; Mudge, Stephen R.; Rae, Anne L. ...
Plant and soil,
01/2003, Letnik:
248, Številka:
1/2
Journal Article
Recenzirano
Transport of inorganic phosphate (Pi) through plant membranes is mediated by a number of families of transporter proteins. Studies on the topology, function, regulation and sites of expression of the ...genes that encode the members of these transporter families are enabling roles to be ascribed to each of them. The Pht1 family, of which there are nine members in the Arabidopsis genome, includes proteins involved in the uptake of Pi from the soil solution and the redistribution of Pi within the plant. Members of this family are H2PO4-/H+ symporters. Most of the genes of the Pht1 family that are expressed in roots are up-regulated in P-stressed plants. Two members of the Pht1 family have been isolated from the cluster roots of white lupin. These same genes are expressed in non-cluster roots. The evidence available to date suggests that there are no major differences between the types of transport systems that cluster roots and non-cluster roots use to acquire Pi. Differences in uptake rates between cluster and non-cluster roots can be ascribed to more high-affinity Pi transporters in the plasma membranes of cluster roots, rather than any difference in the characteristics of the transporters. The efficient acquisition of Pi by cluster roots arises primarily from their capacity to increase the availability of soil Pi immediately adjacent to the rootlets by excretion of carboxylates, protons and phosphatases within the cluster. This paper reviews Pi transport processes, concentrating on those mediated by the Pht1 family of transporters, and attempts to relate those processes involved in Pi acquisition to likely Pi transport processes in cluster roots.
Summary
Sustained expression of transgenes in specified developmental patterns is commonly needed in plant biotechnology, but obstructed by transgene silencing. Here, we present a set of gene design ...rules, tested on the silencing‐susceptible beetle luc and bacterial ims genes, expressed in sugarcane. Designs tested independently or in combination included removal of rare codons, removal of RNA instability sequences, blocking of likely endogenous sRNA binding sites and randomization of non‐rare codons. Stable transgene expression analyses, on multiple independent lines per construct, showed greatest improvement from the removal of RNA instability sequences, accompanied by greatly reduced transcript degradation evident in northern blot analysis. We provide a set of motifs that readily can be eliminated concurrently with rare codons and undesired structural features such as repeat sequences, using Gene Designer 2.0 software. These design rules yielded 935‐ and 5‐fold increased expression in transgenic callus, relative to the native luc and ims sequences; and gave sustained expression under the control of sugarcane and heterologous promoters over several years in greenhouse and field trials. The rules can be applied easily with codon usage tables from any plant species, providing a simple and effective means to achieve sustained expression of otherwise silencing‐prone transgenes in plants.
Transgenic tobacco plants were produced that contained single-copy pART54 T-DNA, with a 35S-uidA gene linked to loxP-flanked kanamycin resistance (nptII) and cytosine deaminase (codA) genes. ...Retransformation of these plants with pCrel (containing 35S transcribed cre recombinase and hygromycin (hpt) resistance genes) resulted in excision of the loxP-flanked genes from the genome. Phenotypes of progeny from selfed-retransformed plants confirmed nptII and codA excision and integration of the cre-linked hpt gene. To avoid integration of the hpt gene, and thereby generate plants totally free of marker genes, we attempted to transiently express the cre recombinase. Agrobacterium tumefaciens (pCrel) was cocultivated with leaf discs of two pART54-transformed lines and shoots were regenerated in the absence of hygromycin selection. Nineteen of 773 (0.25%) shoots showed tolerance to 5-fluorocytosine (5-fc) which is converted to the toxic 5-fluorouracil by cytosine deaminase. 5-fc tolerance in six shoots was found to be due to excision of the loxP-flanked region of the pART54 T-DNA. In four of these shoots excision could be attributed to cre expression from integrated pCrel T-DNA, whereas in two shoots excision appeared to be a consequence of transient cre expression from pCrel T-DNA molecules which had been transferred to the plant cells but not integrated into the genome. The absence of selectable marker genes was confirmed by the phenotype of the T(1) progeny. Therefore, through transient cre expression, marker-free transgenic plants were produced without sexual crossing. This approach could be applicable to the elimination of marker genes from transgenic crops which must be vegetatively propagated to maintain their elite genotype.
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