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Tseng M.-N.,National Taiwan University | Tseng M.-N.,Kaohsiung District Agricultural Research and Extension Station | Chung C.-L.,National Taiwan University | Tzean S.-S.,National Taiwan University
PLoS ONE | Year: 2014

The entomopathogenic fungus Metarhizium anisopliae MA05-169 is a transformant strain that has been metabolically engineered to express dihydroxynaphthalene-melanin biosynthesis genes. In contrast to the wild type strain, the transformant displays a greater resistance to environmental stress and a higher virulence toward target insect host. However, the underlying mechanisms for these characteristics remain unclear; hence experiments were initiated to explore the possible mechanism(s) through physiological and molecular approaches. Although both transformant and wild type strains could infect and share the same insect host range, the former germinated faster and produced more appressoria than the latter, both in vivo and in vitro. The transformant showed a significantly shorter median lethal time (LT50) when infecting the diamondback moth (Plutella xylostella) and the striped flea beetle (Phyllotreta striolata ), than the wild type. Additionally, the transformant was more tolerant to reactive oxygen species (ROS), produced 40-fold more orthosporin and notably overexpressed the transcripts of the pathogenicity-relevant hydrolytic enzymes (chitinase, protease, and phospholipase) genes in vivo. In contrast, appressorium turgor pressure and destruxin A content were slightly decreased compared to the wild type. The transformant's high anti-stress tolerance, its high virulence against five important insect pests (cowpea aphid Aphis craccivora, diamondback moth Pl. xylostella, striped flea beetle Ph. striolata, and silverleaf whitefly Bemisia argentifolii) and its capacity to colonize the root system are key properties for its potential bio-control field application. © 2014 Tseng et al.


Lin Y.-H.,Kaohsiung District Agricultural Research and Extension Station
African Journal of Biotechnology | Year: 2010

Aluminum (Al) is a biotoxic which often influences the absorption of nutrients by plants in strongly acidic soils. In this experiment, the effect of Al on root growth, absorption of macronutrients; phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg) and micronutrients; iron (Fe), manganese (Mn), copper (Cu) and zinc (Zn) by two pineapple cultivars (ig. Al-resistant Cayenne and Alsensitive Tainung No.17) were studied. Four levels of Al concentration treatments were imposed to the hydroponic solutions containing 0, 100, 200 and 300 μM AlCl3. After treatments with Al for four weeks, the root elongation of Cayeen was increased with Al concentration, however, Tainung No.17 was decreased. The dry weight of Cayenne and Tainung No.17 was increased and decreased with Al concentration, respectively. The absorptions of macronutrients and micronutrients were not affected in Al-resistant Cayenne. However, the absorption of Ca, Mg and K were inhibited when AlCl3 was 200 μM, and Fe, Mn and Cu absorption were inhibited significantly when AlCl3 was 300 μM in the Al-sensitive Tainung No.17. On the other hand, Ca, Mg and K absorption increased significantly in Cayenne when AlCl3 was 200 μM. It is possible that Ca, Mg and K uptake was an important clue for Al-resistant pineapple to resist Al in root. © 2010 Academic Journals.


Lin Y.,Kaohsiung District Agricultural Research and Extension Station
Clean - Soil, Air, Water | Year: 2010

The application of potassium fertilizer by farmers is often not appropriate and causing environmental pollution. By understanding the adsorptive characteristics of potassium (K) on different soils, we can prevent excessive application of K-fertilizer that can cause environmental impact. The Gapon exchange coefficient (K G), for exchange between K and Ca, was considered as an important factor influencing the adsorption of K. This study was conducted to compare the constant K G of five important farm soils in Taiwan. The K G and CEC were then used to predict K buffering capacity (PBC). Finally, the relationship between exchangeable K ratio (E P) and K adsorption ratio (PAR) was examined. The results show that five soils have same trends, indicating that K G decreases with increase in K saturation. The CEC and K G of Liuying (Ly) soil are both high, so that their K buffering capacity is high. The K G and CEC of Chanjing (Cj) and Sanhua (Sh) soil show moderate values. The CEC of Erling (El) soil is high, but its K G is low, so that its K buffering capacity is moderate. On the other hand, the K G of Newniaokang (Nnk) soil is high but its CEC is low, so its K buffering capacity is also moderate. The correlations between E P and PAR of five soil show linear relationship at three treatments of CaCl 2 concentration. This study may provide an important clue to the fertilization management of K-fertilizer on the different soil properties in Taiwan.This paper describes on the basis of simple equations exchange behaviors between potassium and other cations in soil. Thus, it may provide a clue to the application method of K-fertilizers. © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Lin Y.-H.,Kaohsiung District Agricultural Research and Extension Station | Su P.,Kaohsiung District Agricultural Research and Extension Station
Clean - Soil, Air, Water | Year: 2010

Humic acid plays an important role in the distribution of heavy metals in the environment. The aims of this study were conducted to evaluate the spectroscopic characteristics and aluminum (Al) adsorption of humic acids which were extracted from four composts. The functional groups were determined by Fourier transform infrared spectroscopy (FTIR) and solid-state 13C nuclear magnetic resonance spectroscopy (13C-NMR). The results showed that the aromatic groups were all found in the humic acids of the four composts, and the surface of humic acids included carboxylic group, hydroxylic group, and amino group. The experiment of Al adsorption was described by Freundlich equation. It showed that the adsorption of Al by humic acid from compost of cattle manure was higher, but that from the compost of pig manure, lemon manure, tea manure in equal preparation was lower. The adsorptive behavior was different due to the interaction by functional groups (-OH and -COOH) with Al. These results can describe the fate and transportation of Al in the soil of different organic fertilizer. The adsorption capacity of well-matured composts for Al ions differs due to humic acid content. Application of such composts to soils and their effects on the fate and transport of Al ions are described here. Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Tseng M.N.,National Taiwan University | Tseng M.N.,Kaohsiung District Agricultural Research and Extension Station | Chung P.C.,National Taiwan University | Chung P.C.,Miaoli District Agricultural Research and Extension Station | Tzean S.S.,National Taiwan University
Applied and Environmental Microbiology | Year: 2011

Entomopathogenic fungi have been used for biocontrol of insect pests for many decades. However, the efficacy of such fungi in field trials is often inconsistent, mainly due to environmental stresses, such as UV radiation, temperature extremes, and desiccation. To circumvent these hurdles, metabolic engineering of dihydroxynaphthalene (DHN) melanin biosynthetic genes (polyketide synthase, scytalone dehydratase, and 1,3,8-trihydroxynaphthalene reductase genes) cloned from Alternaria alternata were transformed into the amelanotic entomopathogenic fungus Metarhizium anisopliae via Agrobacterium-mediated transformation. Melanin expression in the transformant of M. anisopliae was verified by spectrophotometric methods, liquid chromatography/mass spectrometry (LC/MS), and confocal microscopy. The transformant, especially under stresses, showed notably enhanced antistress capacity and virulence, in terms of germination and survival rate, infectivity, and reduced median time to death (LT 50) in killing diamondback moth (Plutella xylostella) larvae compared with the wild type. The possible mechanisms in enhancing the stress tolerance and virulence, and the significance and potential for engineering melanin biosynthesis genes in other biocontrol agents and crops to improve antistress fitness are discussed. © 2011, American Society for Microbiology.


Liao P.-C.,National Pingtung University of Science and Technology | Tsai C.-C.,Kaohsiung District Agricultural Research and Extension Station | Chou C.-H.,China Medical University at Taichung | Chiang Y.-C.,National Sun Yat - sen University
International Journal of Molecular Sciences | Year: 2012

The landrace strains of Momordica charantia are widely cultivated vegetables throughout the tropics and subtropics, but not in Taiwan, a continental island in Southeast Asia, until a few hundred years ago. In contrast, the related wild populations with smaller fruit sizes are native to Taiwan. Because of the introduction of cultivars for agricultural purposes, these two accessions currently exhibit a sympatric or parapatric distribution in Taiwan. In this study, the cultivars and wild samples from Taiwan, India, and Korea were collected for testing of their hybridization and evolutionary patterns. The cpDNA marker showed a clear distinction between accessions of cultivars and wild populations of Taiwan and a long divergence time. In contrast, an analysis of eight selectively neutral nuclear microsatellite loci did not reveal a difference between the genetic structures of these two accessions. A relatively short divergence time and frequent but asymmetric gene flows were estimated based on the isolation-with-migration model. Historical and current introgression from cultivars to wild populations of Taiwan was also inferred using MIGRATE-n and BayesAss analyses. Our results showed that these two accessions shared abundant common ancestral polymorphisms, and the timing of the divergence and colonization of the Taiwanese wild populations is consistent with the geohistory of the Taiwan Strait land bridge of the Last Glacial Maximum (LGM). Long-term and recurrent introgression between accessions indicated the asymmetric capacity to receive foreign genes from other accessions. The modern introduction of cultivars of M. charantia during the colonization of Taiwan by the Han Chinese ethnic group enhanced the rate of gene replacement in the native populations and resulted in the loss of native genes. © 2012 by the authors; licensee MDPI, Basel, Switzerland.


Tsai C.-C.,Kaohsiung District Agricultural Research and Extension Station
Acta Horticulturae | Year: 2014

Sixteen polymorphic microsatellite loci were developed for mango (Mangifera indica L.), to evaluate genetic diversity and identify cultivars of this important commercial fruit tree in East Asia. The 16 polymorphic microsatellite markers were developed using a magnetic bead-enrichment method and polymorphisms were identified when DNA samples from 22 mango cultivars were screened. The number of alleles ranged from 2 to 8, with an expected heterozygosity ranging from 0.304 to 0.826. The polymorphism information content (PIC) ranged from 0.253 to 0.756 (average = 0.543). These polymorphic loci should be useful and convenient for further studies of the genetic diversity and identification of cultivars in mango.


Tsai C.C.,Kaohsiung District Agricultural Research and Extension Station
Acta Horticulturae | Year: 2014

Phalaenopsis is one of the important economic orchids in the world. To date, thousands of Phalaenopsis cultivars have been bred and commercialized. In order to introduce unique characteristics of the spur structure from the flower of Amesiella into Phalaenopsis germplasm, intergeneric hybridization between Phalaenopsis 'Sogo Yukidian' and Amesiella philippinensis were conducted by artificial pollination. Four months later, the capsules were harvested. The immature embryos with placenta were manipulated and placed into germination medium. Approximately 300 intergeneric embryos of each capsule were rescued. After subculturing for one year in the culture medium and two-week hardiness, intergeneric hybrids were cultivated in greenhouse. After one and half years of cultivation, the first flower of the hybrid bloomed. The new hybrid was named as Amenopsis 'Kaohsiung Magic' and registered at the Royal Horticulture Society. Leaf morphology of the hybrid is lanceolate, coriaceus and dark green. The hybrid bears greenish-white flower with black markings on the lip and a short spur at the back of the lip.


Chen J.-H.,National Chung Hsing University | Lin Y.-H.,Kaohsiung District Agricultural Research and Extension Station
Soil Science and Plant Nutrition | Year: 2010

Aluminum (Al) is biotoxic, often active in acid soil and retards the growth of crop roots. Cayenne is a type of pineapple cultivar that can be well cultivated in a strongly acid environment containing AlCl3 concentrations up to 300 μmol L-1. In addition to organic acids, variations in the proteins in root apices are regarded as the mechanism involved in Al resistance. The objective of the present study was to explore the responsive proteins of Al stress in a known Al-resistant pineapple cultivar (i.e. Cayenne). After root emergence, pineapple seedlings were exposed to hydroponic solutions each containing 0 and 300 μmol L-1 AlCl3 for 4 weeks. The total proteins in the root apices were separated using 2-D electrophoresis and a total of 17 apparently differential spots were identified by mass spectrometry, with 10 upregulated and seven downregulated proteins. The root apices of Cayenne under Al stress could be characterized by cellular activities involved for example, carbohydrate metabolism, organic acid production, energy metabolism, alleviating redox damage and root phenotypical change, which are critical for plant survival under Al toxicity. In contrast, there are one hypothetical and three unknown proteins that play unknown roles in Al resistance and warrant further investigation. The present study may provide an important clue to future proteomic research on Al-resistant mechanisms in pineapple. © 2010 Japanese Society of Soil Science and Plant Nutrition.


Patent
Kaohsiung District Agricultural Research And Extension Station and National Taiwan University | Date: 2010-11-26

The present invention cloned melanin biosynthesis genes encoding polyketide synthase (PKS), scytalone dehydratase (SCD) and 1.3.8-trihydroxynaphthalene reductase (THN) from the dematiaceous Alternasia alternate into plasmid pCAMBIA1300, followed by transformation of the plasmid into Matarhizium anisopliae via Agrobacterium tumefaciens-mediated transformation. The transformant was able to express the abovementioned genes and synthesize melanin, which then showed enhanced UV tolerance. The transcription and expression of these melanin genes were confirmed in several pathways. The tolerances toward UV radiation, drought and high temperature were increased significantly in these transformants. In addition, the host insects were more susceptible to these transformants under UV radiation.

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