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This research aims to investigate the potential of utilizing pomegranate peel powder (PPP) as a natural preservative in muffin preparation. Pomegranate peel is a rich source of bioactive compounds, including phenolics, flavonoids, and tannins, which possess high antioxidant and antimicrobial properties. The In-Vitro antifungal activity of pomegranate peel powder (8% PPP), potassium sorbate (0.1% PS) and calcium propionate (0.5% CP) was assessed against Penicillium sp. and Aspergillus sp. using poison food technique. The PPP showed the anti-fungal activity by delaying the growth of microorganism on media plate similar to the PS and CP. The effect of utilization of PPP on quality characteristics of muffins were compared with the muffins with chemical preservatives (0.1% PS and 0.5% CP). The viscosity and specific gravity of batter significantly increased from 7.98 to 11.87 Pa s and 1.089-1.398 respectively on addition of 8% PPP. The optical microscopic structure of PPP added batter revealed the decrease in the number of air cells from 24 to 12 with radius range of 6.42-72.72 µm and area range of 511.03-15,383.17 µm2. The functional properties of flour with PPP had higher water absorption capacity, foaming stability, emulsification activity and emulsion stability than others. The addition of PPP significantly increase the weight (32.83 g), and decrease the height (31.3 mm), volume (61.43 cm3), specific volume (1.67 cm3/g) and baking loss (10.19%). The 418.36% increase in fibre content, 14.46% and 18.46% decrease in carbohydrates and energy value was observed in muffin with 8% PPP as compared to control respectively. The total phenols was increased from 0.92 to 12.5 mg GAE/100 g, total tannin from 0.2 to 8.27 mg GAE/100 g, In-vitro antioxidant activity by DPPH from 6.97 to 29.34% and In-vitro antioxidant activity by FRAP from 0.497 to 2.934 mg AAE/100 g in muffins added with 8% PPP. The muffin with PPP was softer than control and muffin with 0.1% PS. The addition of PPP resulted to improve in muffin texture but taste slightly bitter. During the storage of muffins at room temperature (27-30 °C), the moisture content of muffin with PPP was reduced from 17.04 to 13.23% which was higher than the rest of the treatments. Similarly, the hardness of sample with PPP was higher than the sample with 0.5% CP, but lowers than control and sample with 0.1% PS throughout the storage period. The results suggest that pomegranate peel powder can be successfully used as a natural preservative in place of chemical preservatives in muffins, to extend the shelf life. This study provides the opportunity to use PPP as functional ingredient and natural preservative in different bakery products.
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Conservación de Alimentos , Conservantes de Alimentos , Granada (Fruta) , Polvos , Conservantes de Alimentos/farmacología , Conservantes de Alimentos/química , Granada (Fruta)/química , Conservación de Alimentos/métodos , Penicillium/efectos de los fármacos , Antioxidantes/farmacología , Antioxidantes/química , Antifúngicos/farmacología , Antifúngicos/química , Aspergillus/efectos de los fármacos , Aspergillus/crecimiento & desarrollo , Frutas/química , Almacenamiento de Alimentos/métodos , Extractos Vegetales/farmacología , Extractos Vegetales/químicaRESUMEN
PURPOSE OF THE REVIEW: Pomegranate is one of the super fruit and a storehouse of several antioxidants and health-promoting compounds which can act as a natural food additive. The pomegranate processing industry generates huge quantities of by-products, particularly peels (50% of fresh fruit weight), that cause environmental pollution due to improper disposal. In this perspective, the present review article focuses on the chemical composition of pomegranate peel and its application as a natural food additive in different food products such as bakery, dairy, meat/meat products, fish/fish products, edible oils, and packaging materials. RECENT FINDINGS: There is a continuous demand for processed foods exhibiting natural food additives over foods containing synthetic additives/colorants, which can cause serious health implications such as cancer with regular consumption. The food industry is looking for an alternative to synthetic/artificial food additives. To overcome these problems, pomegranate peel or its extract can be used as a natural biopreservative in food products that are prone to fat oxidation and microbial growth. Pomegranate peel contains bioactive compounds, especially tannins, phenolic acids, and flavonoids, which have nutraceutical value and possess higher antioxidant activity and antimicrobial properties. Due to these properties, pomegranate peel prevents lipid oxidation in fatty foods and can also retard the microbial growth.
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Aditivos Alimentarios , Granada (Fruta) , Animales , Humanos , Aditivos Alimentarios/análisis , Granada (Fruta)/química , Extractos Vegetales/farmacología , Antioxidantes/farmacología , Frutas/químicaRESUMEN
This study reports genome wide characterization and development of first set of microsatellite markers through in silico analysis of eight sequenced Xanthomonas axonopodis pv. punicae strains available in the public database. SSR survey resulted in identification of ~ 4638 perfect SSRs, with mean marker frequency 901 SSRs/Mb and densitiy of 11,006 bp/Mb aross the eight genomes. Frequency distribution graphs revealed hexa-nucleotide repeats were more prominent fowllowed by tri-, tetra-, di- and penta-nucleotides in the analysed genomes. We desinged 2927 SSR primers that are specific to the strain LMG 859 and ePCR confirmed on seven other Xap genomes. This resulted in identification of 542 informative SSRs that are producing single amplicons, from which 66 primers were successfully validated through wet lab experiments on eight Xap isolates of pomegranate. Furthermore, utility of these SSRs were demostrated by analysing molecular diversity among 22 Xap isolates using 20 Xap_SSR primers. SSRs revealed moderate genetic diversity among Xap isolates (61%) and grouped 11 isolates that are repersenting six different states into one cluster. This proved the earlier evidence of wider spread of ST3 type Xap acoss India using Multi locus Sequence Typing (MLST) technique. In summary, Xap_SSR will serve as powerful genomics tools that would helps in monitoring of population dynamics, taxonomy, epidomology and quarantine aspects in bacterial blight pathogen through development of microsatellite based Multilocus Variable number of Tandem repeat analysis (MLVA) in future. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03209-z.
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High density planting system i.e. accommodating a higher number of plants than routine in a given area is an innovative agro-technology to increase yield and thereby early net returns. Due to conventional wide spacing plantation in Nagpur mandarin (Citrus reticulata Blanco), the land remains unutilized as the plant canopy gradually increases over the years. In the present study, Nagpur mandarin (Citrus reticulata Blanco) budded on Rangpur lime rootstock was evaluated under six different planting spacings. It was observed that the organic carbon (1.10-1.82%) and major nutrients viz. N (309-430 kg ha-1), P (20-54 kg ha-1) and K (291-810 kg ha-1) increased vis-à-vis plant density and was highest under 2 × 2 m spacing. Plants were tallest at 2 × 2 m spacing with the higher PAR interception (88.2) and the lowest leaf area index (1.09). Fruit yield on area basis, under 2 × 2 m spacing was 26, 7.1, 4.6 times more as compared to conventional plantation during the first, second and third year, respectively. At fifth year of crop harvest, the highest B:C ratio (6.36) was recorded in 6 × 3 m followed by 4 × 2 m and 2 × 2 m.
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Children with high body mass index (BMI) are at risk of iron deficiency. In present study, 71 children with overweight or obesity were screened for iron deficiency. Mean BMI, ferritin and plasma soluble transferrin receptor (sTrfR) levels were 26.1 kg/m2, 41.9 µg/L and 0.375 mg/L, respectively. Twenty (28%) children had anemia, and 44 (62%) had an underlying hypoferraemic state.
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Anemia Ferropénica , Sobrepeso , Obesidad Infantil , Adolescente , Anemia Ferropénica/complicaciones , Anemia Ferropénica/epidemiología , Biomarcadores/sangre , Índice de Masa Corporal , Niño , Preescolar , Estudios Transversales , Femenino , Ferritinas/sangre , Humanos , India/epidemiología , Masculino , Sobrepeso/complicaciones , Sobrepeso/epidemiología , Obesidad Infantil/complicaciones , Obesidad Infantil/epidemiologíaRESUMEN
Soluble serum transferrin receptor is derived from erythroid transferrin receptor expressed on surface of developing erythroid cells. It can be detected in blood using sensitive ELISA methodology and blood levels reflect physiological iron dependent erythropoiesis state in bone marrow. Normal adult levels vary from 2 to 5 mg/l. However, pediatric studies are few and describe normal ranges to the tune of 1.0-3.0 mg/l, which are relatively lower than that of adults. In present study 40 healthy children (2-12 years) were evaluated to establish normal soluble transferrin receptor range. The mean transferrin receptor levels were 0.39 mg/l with a range of 0.17-2.1 mg/l. The levels were low as compared to mean levels described in other studies from West and our country (4.39 and 2.0 mg/l respectively). Since, no internationally standard method for reporting and testing for transferrin receptor levels are yet available, hence it is imperative to establish normal control ranges in different population cohorts, especially in pediatric age group, to better interpret their levels in diagnostic context.
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Macrofungi have long been investigated for various scientific purposes including their food and medicinal characteristics. Their role in aerobiology as a fraction of the primary biological aerosol particles (PBAPs), however, has been poorly studied. In this study, we present a source of macrofungi with two different but interdependent objectives: (i) to characterize the macrofungi from a tropical dry evergreen biome in southern India using advanced molecular techniques to enrich the database from this region, and (ii) to assess whether identified species of macrofungi are a potential source of atmospheric PBAPs. From the DNA analysis, we report the diversity of the terrestrial macrofungi from a tropical dry evergreen biome robustly supported by the statistical analyses for diversity conclusions. A total of 113 macrofungal species belonging to 54 genera and 23 families were recorded, with Basidiomycota and Ascomycota constituting 96% and 4% of the species, respectively. The highest species richness was found in the family Agaricaceae (25.3%) followed by Polyporaceae (15.3%) and Marasmiaceae (10.8%). The difference in the distribution of commonly observed macrofungal families over this location was compared with other locations in India (Karnataka, Kerala, Maharashtra, and West Bengal) using two statistical tests. The distributions of the terrestrial macrofungi were distinctly different in each ecosystem. We further attempted to demonstrate the potential role of terrestrial macrofungi as a source of PBAPs in ambient air. In our opinion, the findings from this ecosystem of India will enhance our understanding of the distribution, diversity, ecology, and biological prospects of terrestrial macrofungi as well as their potential to contribute to airborne fungal aerosols.
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Biodiversidad , Ecosistema , Hongos , Plantas/microbiología , Clima Tropical , Microbiología Ambiental , Hongos/clasificación , Hongos/genética , Hongos/ultraestructura , Incidencia , India , Metagenoma , Metagenómica/métodos , Estaciones del Año , Esporas Fúngicas/ultraestructuraRESUMEN
Engelhard titanium silicate, ETS-4, is a promising new adsorbent for size-selective separation of mixtures of small gases, a leading industrially important example of which is methane-nitrogen separation. Single component equilibrium and kinetics of oxygen, nitrogen, and methane adsorption in Na-ETS-4 and cation-exchanged Sr-ETS-4, measured in an earlier study over a wide range of temperatures and pressures, are analyzed in this study. The adsorbent crystals were synthesized and pelletized under pressure (without any binder), thus giving rise to a bidispersed pore structure with controlling resistance in the micropores. Change in equilibrium and kinetics of adsorption of the aforementioned gases in Sr-ETS-4 due to pore shrinkage with progressively increasing dehydration temperature has also been investigated. Differential uptakes have been measured at various levels of adsorbate loading, which has allowed the elucidation of the nature of concentration dependence of micropore diffusivity. Both homogeneous and heterogeneous models are examined on the equilibrium data, while a bidispersed pore diffusion model is able to capture the differential uptakes very well. On the basis of chemical potential gradient as the driving force for diffusion, the impact of isotherm models on the concentration dependence of micropore diffusivity is also analyzed. It is shown that pore tailoring at the molecular scale by dehydration can improve the kinetic selectivity of nitrogen over methane in Sr-ETS-4 to a promising level. The models investigated are evaluated to identify essential details necessary to reliably simulate a methane-nitrogen separation process using the promising new Sr-ETS-4 adsorbent.
RESUMEN
Energetic heterogeneity has been investigated for Engelhard titanium silicate Na-ETS-4 adsorbent and its Sr-exchanged variant, Sr-ETS-4. Na-ETS-4 was nearly homogeneous, while Sr exchange seemed to induce some degree of energetic heterogeneity in the sample, which diminished upon dehydration at higher temperature. Analysis of the entropy change during adsorption showed that the adsorbate molecules at low as well as moderate loading possess entropy greater than that predicted by the 2-D mobile film model, the excess being attributed to vibrational freedom. The wavelength of this vibration decreased with increasing coverage, as expected. For oxygen, the observed entropy drops in Na-ETS-4 and in Sr-ETS-4 are comparable, whereas, for nitrogen and methane, Sr exchange resulted in a greater entropy drop than in Na-ETS-4, suggesting greater restriction to movement in the Sr-exchanged sample. This study presents a simplistic yet effective understanding of the energetic behavior of the adsorbed molecules in ETS-4 adsorbent. This is vital to a thorough energetic characterization and study of the adsorption phenomenon in these new, promising adsorbents.
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Strontium-exchanged titanium silicate, Sr-ETS-4, is a new molecular sieve with promises for exciting applications in gas separation. The literature mentions poor thermal stability and low adsorption capacity as the drawbacks of the as-synthesized Na-ETS-4 and therefore the need for cation exchange. Upon ion exchange, Sr-ETS-4 shows appreciable kinetic selectivity between methane and nitrogen, which can be improved further by controlled dehydration. In this study, we trace the changes at the molecular level behind this improvement. By combining distributed information in the literature, it is shown that the governing factors are changes in pore geometry with progressive dehydration, changes in ion occupancy, and relocation of cations due to Sr exchange.
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Post-transcriptional gene silencing (PTGS) is a sequence-specific RNA degradation mechanism that is widespread in eukaryotic organisms. It is often associated with methylation of the transcribed region of the silenced gene and with accumulation of small RNAs (21 to 25 nucleotides) homologous to the silenced gene. In plants, PTGS can be triggered locally and then spread throughout the organism via a mobile signal that can cross a graft junction. Previously, we showed that the helper component-proteinase (HC-Pro) of plant potyviruses suppresses PTGS. Here, we report that plants in which PTGS has been suppressed by HC-Pro fail to accumulate the small RNAs associated with silencing. However, the transgene locus of these plants remains methylated. Grafting experiments indicate that HC-Pro prevents the plant from responding to the mobile silencing signal but does not eliminate its ability to produce or send the signal. These results demonstrate that HC-Pro functions downstream of transgene methylation and the mobile signal at a step preceding accumulation of the small RNAs.
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Cisteína Endopeptidasas/genética , Silenciador del Gen/fisiología , ARN de Planta/metabolismo , Supresión Genética , Transgenes/fisiología , Proteínas Virales/genética , Algoritmos , Northern Blotting , Southern Blotting , Glucuronidasa/análisis , Glucuronidasa/genética , Técnicas In Vitro , Metilación , Virus de Plantas/genética , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/metabolismo , Plantas Tóxicas , Reacción en Cadena de la Polimerasa , Procesamiento Postranscripcional del ARN , ARN de Planta/genética , Homología de Secuencia de Ácido Nucleico , Transducción de Señal , Nicotiana/genética , Nicotiana/metabolismo , Transcripción Genética , Trasplantes , Proteínas Virales/antagonistas & inhibidoresRESUMEN
Posttranscriptional gene silencing (PTGS) is an ancient eukaryotic regulatory mechanism in which a particular RNA sequence is targeted and destroyed. The helper component-proteinase (HC-Pro) of plant potyviruses suppresses PTGS in plants. Using a yeast two-hybrid system, we identified a calmodulin-related protein (termed rgs-CaM) that interacts with HC-Pro. Here we report that rgs-CaM, like HC-Pro itself, suppresses gene silencing. Our work is the first report identifying a cellular suppressor of PTGS.
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Cisteína Endopeptidasas/metabolismo , Silenciador del Gen , Nicotiana/genética , Proteínas de Plantas/metabolismo , Plantas Tóxicas , Proteínas Virales/metabolismo , Agrobacterium tumefaciens/genética , Secuencia de Aminoácidos , Genes de Plantas , Proteínas Fluorescentes Verdes , Proteínas Luminiscentes/genética , Datos de Secuencia Molecular , Proteínas de Plantas/química , Proteínas de Plantas/genética , Tumores de Planta/genética , Plantas Modificadas Genéticamente , Plásmidos , Potexvirus/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN de Planta/genética , ARN de Planta/metabolismo , Nicotiana/metabolismo , Transcripción Genética , TransgenesRESUMEN
Post-transcriptional gene silencing (PTGS) is a fundamental regulatory mechanism operating in diverse types of organisms, but the cellular components of the gene silencing machinery and the regulation of the process are not understood. Recent findings that cytoplasmically replicating RNA viruses act as both targets and inducers of PTGS has led to the idea that PTGS may have evolved as an anti-viral defense mechanism in plants. Consistent with this hypothesis, it has been found that certain plant viruses encode proteins that suppress PTGS. From a practical standpoint, an understanding of the mechanisms by which viruses regulate PTGS may well lead to better ways to control gene expression in plants. It is often desirable to overexpress selected beneficial genes or to silence detrimental ones in order to confer a particular phenotype. Induction of PTGS using RNA viruses as vectors or as transgenes provides a reliable and efficient way to interfere with the expression of a specific gene or with a family of genes. Conversely, expression of viral suppressors has significant potential to improve yields in technologies that use plants to express beneficial gene products. Given the antiviral nature of gene silencing in plants and the indications that PTGS is an ancient mechanism in eukaryotic organisms, understanding the phenomenon in plants could well lead to the development of anti-viral strategies in both plants and animals.
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Silenciador del Gen , Plantas/genética , Virus ARN/fisiología , Plantas/virología , Procesamiento Postranscripcional del ARN , Virus ARN/genética , ARN Viral/genética , ARN Viral/fisiologíaRESUMEN
Homology-dependent gene silencing is a regulatory mechanism that limits RNA accumulation from affected loci either by suppression of transcription (transcriptional gene silencing, TGS) or by activation of a sequence-specific RNA degradation process (post-transcriptional gene silencing, PTGS). The P1/HC-Pro sequence of plant potyviruses and the 2b gene of the cucumber mosaic virus have been shown to interfere with PTGS. The ability of these viral suppressors of PTGS to interfere with TGS was tested using the 271 locus which imposes TGS on transgenes under 35S or 19S promoters and PTGS on the endogenous nitrite reductase gene (Nii). Both P1/HC-Pro and 2b reversed PTGS of Nii genes in 271-containing tobacco plants, but failed to reverse TGS of 35S-GUS transgenes in the same plant. P1/HC-Pro expression from a transgene also failed to suppress either the initiation or maintenance of TGS imposed by the NOSpro-silencing locus, H2. These results indicate that PTGS and TGS operate through unlinked pathways or that P1/HC-Pro and 2b interfere at step(s) in PTGS that are downstream of any common components in the two pathways. The data suggest a simple assay to identify post-transcriptionally silenced transgenic lines with the potential to be stably converted to high expressing lines.
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Silenciador del Gen , Genes Virales , Virus de Plantas/genética , Supresión Genética , Cucumovirus/genética , Plantas Modificadas Genéticamente , Plantas Tóxicas , Potyvirus/genética , Nicotiana , Transcripción GenéticaRESUMEN
Chlamydia are obligate intracellular eubacteria that are phylogenetically separated from other bacterial divisions. C. trachomatis and C. pneumoniae are both pathogens of humans but differ in their tissue tropism and spectrum of diseases. C. pneumoniae is a newly recognized species of Chlamydia that is a natural pathogen of humans, and causes pneumonia and bronchitis. In the United States, approximately 10% of pneumonia cases and 5% of bronchitis cases are attributed to C. pneumoniae infection. Chronic disease may result following respiratory-acquired infection, such as reactive airway disease, adult-onset asthma and potentially lung cancer. In addition, C. pneumoniae infection has been associated with atherosclerosis. C. trachomatis infection causes trachoma, an ocular infection that leads to blindness, and sexually transmitted diseases such as pelvic inflammatory disease, chronic pelvic pain, ectopic pregnancy and epididymitis. Although relatively little is known about C. trachomatis biology, even less is known concerning C. pneumoniae. Comparison of the C. pneumoniae genome with the C. trachomatis genome will provide an understanding of the common biological processes required for infection and survival in mammalian cells. Genomic differences are implicated in the unique properties that differentiate the two species in disease spectrum. Analysis of the 1,230,230-nt C. pneumoniae genome revealed 214 protein-coding sequences not found in C. trachomatis, most without homologues to other known sequences. Prominent comparative findings include expansion of a novel family of 21 sequence-variant outer-membrane proteins, conservation of a type-III secretion virulence system, three serine/threonine protein kinases and a pair of parologous phospholipase-D-like proteins, additional purine and biotin biosynthetic capability, a homologue for aromatic amino acid (tryptophan) hydroxylase and the loss of tryptophan biosynthesis genes.
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Proteínas Bacterianas/genética , Chlamydia trachomatis/genética , Chlamydophila pneumoniae/genética , Genoma Bacteriano , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Chlamydia trachomatis/metabolismo , Chlamydia trachomatis/patogenicidad , Chlamydophila pneumoniae/metabolismo , Chlamydophila pneumoniae/patogenicidad , Secuencia Conservada , Enzimas/genética , Enzimas/metabolismo , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Datos de Secuencia Molecular , Operón , Homología de Secuencia de Aminoácido , Triptófano/biosíntesisRESUMEN
Analysis of the 1,042,519-base pair Chlamydia trachomatis genome revealed unexpected features related to the complex biology of chlamydiae. Although chlamydiae lack many biosynthetic capabilities, they retain functions for performing key steps and interconversions of metabolites obtained from their mammalian host cells. Numerous potential virulence-associated proteins also were characterized. Several eukaryotic chromatin-associated domain proteins were identified, suggesting a eukaryotic-like mechanism for chlamydial nucleoid condensation and decondensation. The phylogenetic mosaic of chlamydial genes, including a large number of genes with phylogenetic origins from eukaryotes, implies a complex evolution for adaptation to obligate intracellular parasitism.
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Chlamydia trachomatis/genética , Genoma Bacteriano , Análisis de Secuencia de ADN , Aerobiosis , Secuencia de Aminoácidos , Aminoácidos/biosíntesis , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Evolución Biológica , Chlamydia trachomatis/clasificación , Chlamydia trachomatis/metabolismo , Chlamydia trachomatis/fisiología , Reparación del ADN , Metabolismo Energético , Enzimas/química , Enzimas/genética , Humanos , Lípidos/biosíntesis , Datos de Secuencia Molecular , Peptidoglicano/biosíntesis , Peptidoglicano/genética , Filogenia , Biosíntesis de Proteínas , Recombinación Genética , Transcripción Genética , Transformación Bacteriana , VirulenciaRESUMEN
Gene silencing is an important but little understood regulatory mechanism in plants. Here we report that a viral sequence, initially identified as a mediator of synergistic viral disease, acts to suppress the establishment of both transgene-induced and virus-induced posttranscriptional gene silencing. The viral suppressor of silencing comprises the 5'-proximal region of the tobacco etch potyviral genomic RNA encoding P1, helper component-proteinase (HC-Pro) and a small part of P3, and is termed the P1/HC-Pro sequence. A reversal of silencing assay was used to assess the effect of the P1/HC-Pro sequence on transgenic tobacco plants (line T4) that are posttranscriptionally silenced for the uidA reporter gene. Silencing was lifted in offspring of T4 crosses with four independent transgenic lines expressing P1/HC-Pro, but not in offspring of control crosses. Viral vectors were used to assess the effect of P1/HC-Pro expression on virus-induced gene silencing (VIGS). The ability of a potato virus X vector expressing green fluorescent protein to induce silencing of a green fluorescent protein transgene was eliminated or greatly reduced when P1/HC-Pro was expressed from the same vector or from coinfecting potato virus X vectors. Expression of the HC-Pro coding sequence alone was sufficient to suppress virus-induced gene silencing, and the HC-Pro protein product was required for the suppression. This discovery points to the role of gene silencing as a natural antiviral defense system in plants and offers different approaches to elucidate the molecular basis of gene silencing.
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The potentials and limitations of negative-selection systems based on the human herpes simplex virus thymidine kinase type-1 (HSVtk) gene, which causes sensitivity to the nucleoside analog ganciclovir, were examined in tobacco as a model system. There were great differences between individual HSVtk(+) transgenic plants in ganciclovir sensitivity. Inhibition of growth while under selection correlated with HSVtk-tianscnpt levels. Negative selection against HSVtk(+) transformants at the level of Agrobacterium-mediated transformation using a ganciclo-vir/kanamycin double-selection medium (the positive selection marker neomycin phosphotransferase-II gene was in the transformation vector) resulted in a three- to six-fold reduction in the frequency of kanamycin-resistant shoots. The efficiency of negative selection in this case was limited due to the great variation in HSVtk expression, i.e., the frequently occurring transformants with low, or no, ganciclovir sensitivity escaping negative selection. Two independently constructed HSVtk genes showed the same variability of the phenotype in Nicotiana tabacum transformants. Distinct phenotypes, ranging from no regeneration through abnormal or delayed regeneration, were observed when leaf segments were placed on shoot-inducing medium supplemented with 10(-6)-10(-3) M ganciclovir. The highest HSVtk mRNA and ganciclovir sensitivity levels were observed in plants which were transformed with the pSLJ882 chimeric construct. The pSLJ882 plant expression vector carried the coding sequence of HSVtk, whereas plasmid pCX305.1 carried an HSVtk construct retaining the untranslated 5 leader and viral 3 regions. The pCX305.1 transformants showed, at most, a delayed formation of shoots with thin stems and very narrow leaves. Ganciclovir sensitivity showed typical Mendelian segregation. A gene-dosage effect was also seen at the seedling level in the progeny of two transgenic lines.