Development of tissue culture and virus-induced gene silencing for Calendula officinalis

Calendula officinalis is grown widely as an ornamental plant across Europe. It belongs to the large. Asteraceae family. In this study, the aim was to explore the possibilities to use Calendula officinalis as a new model organism for flower development and secondary mechanism studies in Asteraceae. Tissue culture of Calendula officinalis was established using nine different cultivars. Murashige & Skoog (MS) medium with four different combinations of plant growth regulators were tested. Of all these combinations, the medium containing 1mg/l BAP, 0.1 mg/l IAA, and 1mg/l Zeatin achieved highest frequency of adventitious shoot regeneration from hypocotyl and cotyledon explants. Virus-induced gene silencing is a recent developed genetic tool for charactering the gene functions in plants, and extends the range of host plants that are not accessible for Agrobacterium transformation. Here, tobacco rattle virus (TRV)-based VIGS technique was tested in calendula (cv. Single Orange). We used TRV carrying Gerbera hybrid phytoene desaturase (PDS) gene fragment to induce PDS silencing in calendula. Vacuum infiltration and syringe infiltration methods both resulted in photo-bleaching phenotypes in leaves, bracts and petals. Loss-of-function phenotypes occurred on calendula 13 days post-infiltration. In conclusion, the data indicates that calendula explants can be regenerated through tissue culture which is a prerequisite for development of stable transformation methods. However, further optimization is still needed to improve the frequency. In addition, VIGS was applied to silence PDS marker gene expression indicating that this method has potential for gene functional studies in future.

Snoring and sleep apnea in children

Snoring is a primary and major clinical symptom of upper airway obstruction during sleep. Sleep-disordered breathing ranges from primary snoring to significant partial upper airway obstruction, and obstructive sleep apnea. Adult snoring and obstructive sleep apnea have been extensively studied, whereas less is known about these disorders in children. Snoring and more severe obstructive sleep apnea have been shown to have a harmful effect on the neurobehavioral development of children, but the mechanisms of this effect remains unknown. Furthermore, the correlation of this effect to objective sleep study parameters remains poor. This study evaluated the prevalence of snoring in preschool-aged children in Finland. Host and environmental risk factors, and neurobehavioral and neurocognitive symptoms of children suffering from snoring or obstructive sleep apnea were also investigated. The feasibility of acoustic rhinometry in young children was assessed. The prevalence and risk factors of snoring (I) were evaluated by a questionnaire. The random sample included 2100 children aged 1-6 years living in Helsinki. All 3- to 6-year-old children whose parents reported their child to snore always, often, or sometimes were categorized as snorers, and invited to participate to the clinical study (II-IV). Non-snoring children whose parents were willing to participate in the clinical study were invited to serve as controls. Children underwent a clinical ear-nose-throat examination. Emotional, behavioral, and cognitive performances were evaluated by Child Behavioral Checklist (CBCL), Wechsler Preschool and Primary Scale of Intelligence (WPPSI-R) and NEPSY-A Developmental Neuropsychological Assessment (NEPSY). Nasal volume was measured by acoustic rhinometry, and nasal resistance by rhinomanometry. Lateral and posteroanterior cephalometry were performed. A standard overnight ambulatory polysomnography was performed in the home environment. Twenty-six healthy children were tested in order to assess the feasibility of acoustic rhinometry in young children (V). Snoring was common in children; 6.3% of children snored always or often, whereas 81.3% snored never or occasionally. No differences were apparent between snorers and non-snorers regarding age, or gender. Pediatric snoring was associated with recurrent upper respiratory infections, otitis media, and allergic rhinitis. Exposure to parental tobacco smoke, especially maternal smoking, was more common among snorers. Rhinitis was more common among children who exposured to tobacco smoke. Overnight polysomnography (PSG) was performed on 87 children; 74% showed no signs of significant upper airway obstruction during sleep. Three children had obstructive apnea/hypopnea index (OAHI) greater than 5/h. Age, gender, or a previous adenoidectomy or tonsillectomy did not correlate with OAHI, whereas tonsillar size did correlate with OAHI. Relative body weight and obesity correlated with none of the PSG parameters. In cephalometry, no clear differences or correlations were found in PSG parameters or between snorers and non-snorers. No correlations were observed between acoustic rhinometry, rhinomanometry, and PSG parameters. Psychiatric symptoms were more frequent in the snoring group than in the nonsnoring group. In particular, anxious and depressed symptoms were more prevalent in the snoring group. Snoring children frequently scored lower in language functions. However, PSG parameters correlated poorly with neurocognitive test results in these children. This study and previous studies indicate that snoring without episodes of obstructive apnea or SpO2 desaturations may cause impairment in behavioral and neurocognitive functions. The mechanism of action remains unknown. Exposure to parental tobacco smoke is more common among snorers than non-snorers, emphasizing the importance of a smoke-free environment. Children tolerated acoustic rhinometry measurements well.

Nupoutuskivun vaikutus vasikan hyvinvointiin

Nupouttamisella tarkoitetaan vasikan sarvenaiheiden tuhoamista sarvien kasvun estämiseksi esimerkiksi kuumapolton avulla. Nykyisin lähes kaikki Suomessa syntyvistä vasikoista nupoutetaan turvallisuusnäkökohtien vuoksi. Suomen eläinlääkintölainsäädännön mukaan riittävät tiedot ja taidot toimenpiteen suorittamiseksi omaava henkilö saa nupouttaa alle neliviikkoisen vasikan. Eläinlääkäri voi jättää tilalle pistoksena tai suun kautta annosteltavaa tulehduskipulääkettä nupoutuskivun hoitoa varten. Nupouttamisen on useissa tutkimuksissa todettu aiheuttavan vasikalle kipua, joka ilmenee muun muassa pakenemisyrityksinä toimenpiteen yhteydessä sekä levottomuutena, sosiaalisen käyttäytymisen vähenemisenä, kohonneena sydämen sykkeenä ja stressihormonien tason nousuna nupoutuksen jälkeen. Tähänastisissa tutkimuksissa nupoutuskipua on tutkittu pisimmillään kahden vuorokauden ajan. Koska kuumapoltolla nupouttaminen saa aikaan syvän palovamman, voidaan olettaa vasikan kokevan useiden päivien ajan kipua, joka vaikuttaa hyvinvointia alentavasti. Nupoutuksen vaikutusta vasikoiden makuukäyttäytymiseen tutkittiin viiden päivän ajan nupoutustoimenpiteen jälkeen olettaen, että useamman päivän mittainen tulehduskipulääkekuuri parantaisi vasikoiden hyvinvointia. Tutkimukseen valittiin satunnaisesti kymmenen sonnivasikkaa (keski-ikä 27± 5,6 vrk). Vasikat satunnaistettiin kahteen hoitoryhmään, joista toiset saivat nupoutuspäivänä ja sen jälkeen ketoprofeenivalmistetta suun kautta yhteensä viiden päivän ajan, ja toiset vastaavasti lumelääkettä. Kaikki vasikat nupoutettiin rauhoitettuina ja paikallispuudutettuina kaasukuumenteista nupoutuskolvia käyttäen. Vasikoita videoitiin kuuden vuorokauden ajan, alkaen vuorokausi ennen nupoutusta. Videoaineisto analysoitiin Cowlog -tietokoneohjelmalla. Vasikoita lääkinneet henkilöt sekä videoiden analysoinnin suorittaja eivät tienneet, kumpaan hoitoryhmään kukin vasikka kuului. Koetilanne oli siis kaksoissokkoutettu. Nupoutuspäivää ei otettu huomioon tuloksissa, koska vasikat nupoutettiin rauhoitettuna. Nupoutustoimenpide lyhensi vasikoiden keskimääräistä yhteenlaskettua makuuaikaa vuorokaudessa 1028,2 ± 23,8 minuutista (päivä ennen nupoutusta) 886,1 ± 23,8 minuuttiin (nupoutuspäivää seuraava päivä). Nupoutuspäivää seuraavana päivänä vasikoiden keskimääräinen kokonaismakuuaika oli tilastollisesti merkittävästi lyhyempi verrattuna kaikkiin muihin päiviin (p<0,05). Tuloksissa oli havaittavissa tilastollinen tendenssi, jonka mukaan lumelääkittyjen vasikoiden makuujaksojen lukumäärä vuorokaudessa (22,7±1,6 kertaa) oli tulehduskipulääkittyihin vasikoihin (17,7±1,6 kertaa) verrattuna suurempi koko seurantajakson aikana (p=0,06). Tulehduskipulääkittyjen vasikoiden makuujaksojen keskimääräinen pituus koko seurantajakson aikana oli merkittävästi suurempi (57,3 ± 3,1 minuuttia) kuin lumelääkittyjen vasikoiden (45,0 ± 3,1 minuuttia) (p=0,02). Nupoutus lyhensi kaikkien vasikoiden päivittäistä kokonaismakuuaikaa, mikä viittaa vasikoiden kokeneen akuuttia kipua tulehduskipulääkityksestä huolimatta. Akuutti kipu ja stressi ilmenivät levottomuutena vasikoiden makuukäyttäytymisessä. Riittävä ja laadukas uni on edellytys vasikan kasvulle ja hyvinvoinnille, mikä tekee aiheesta tärkeän myös taloudellisesta näkökulmasta. Tuloksien perusteella voidaan todeta, että nupoutuksen aiheuttama kipu kestää useita päiviä, mutta jatkotutkimuksia tarvitaan kivunhoitovaihtoehtojen osalta sekä kivun kestosta suuremmalla otannalla.

A Parallel Progressive Refinement Image Rendering Algorithm on a Scalable Multithreaded VLSI Processor Array

In this paper we develop a multithreaded VLSI processor linear array architecture to render complex environments based on the radiosity approach. The processing elements are identical and multithreaded. They work in Single Program Multiple Data (SPMD) mode. A new algorithm to do the radiosity computations based on the progressive refinement approach[2] is proposed. Simulation results indicate that the architecture is latency tolerant and scalable. It is shown that a linear array of 128 uni-threaded processing elements sustains a throughput close to 0.4 million patches/sec.

De "de" : Estudio histórico-comparativo de los usos y la semántica de la preposición "de" en español

El presente estudio supone un intento de describir y analizar el uso de la preposición "de" sobre la base de un corpus diacrónico, con énfasis en las diferentes relaciones semánticas que establece. Partiendo de un total de más de 16.000 casos de "de" hemos establecido 48 categorías de uso, que corresponden a cuatro tipos de construcción sintáctica, a saber, el uso de "de" como complemento de nombres (CN), verbos (CV), adjetivos (CA) y, finalmente, su uso como núcleo de expresiones adverbiales independientes (CI). El estudio consta de tres partes fundamentales. En la parte I, se introduce la Lingüística Cognitiva, que constituye la base teórica esencial del trabajo. Más exactamente, se introducen conceptos como la teoría del prototipo, la teoría de las metáforas conceptuales y la gramática cognitiva, especialmente las ideas de "punto de referencia" y "relación intrínseca" (Langacker 1995, 1999). La parte II incluye el análisis de las 48 categorías. En esta parte se presentan y comentan casi 2.000 ejemplos del uso contextual de "de" extraídos del corpus diacrónico. Los resultados más importantes del análisis pueden resumirse en los siguientes puntos: El uso de "de" sigue siendo esencialmente el mismo en la actualidad que hace 800 años, en el sentido de que todas las 48 categorías se identifican en todas las épocas del corpus. El uso de "de" como complemento nominal va aumentando, al contrario de lo que ocurre con su uso como complemento verbal. En el contexto nominal son especialmente las relaciones posesivas más abstractas las que se hacen más frecuentes, mientras que en el contexto verbal las relaciones que se hacen menos frecuentes son las de separación/alejamiento, causa, agente y partitivo indefinido. Destaca la importancia del siglo XVIII como época de transición entre un primer estado de las cosas y otro posterior, en especial en relación con el carácter cada vez más abstracto de las relaciones posesivas así como con la disminución de las categorías adverbales de causa, agente y partitivo. Pese a la variación en el contexto inmediato de uso, el núcleo semántico de "de" se mantiene inalterado. La parte III toma como punto de partida los resultados del análisis de la parte II, tratando de deslindar el aporte semántico de la preposición "de" a su contexto de uso del valor de la relación en conjunto. Así, recurriendo a la metodología para determinar el significado básico y la metodología para determinar lo que constituyen significados distintos de una preposición (Tyler , Evans 2003a, 2003b), se llega a la hipótesis de que "de" posee cuatro significados básicos, a saber, 'punto de partida', 'tema/asunto', 'parte/todo' y 'posesión'. Esta hipótesis, basada en las metodologías de Tyler y Evans y en los resultados del análisis de corpus, se intenta verificar empíricamente mediante el uso de dos cuestionarios destinados a averiguar hasta qué punto las distinciones semánticas a las que se llega por vía teórica son reconocidas por los hablantes nativos de la lengua (cf. Raukko 2003). El resultado conjunto de los dos acercamientos tanto refuerza como especifica la hipótesis. Los datos que arroja el análisis de los cuestionarios parecen reforzar la idea de que el núcleo semántico de "de" es complejo, constando de los cuatro valores mencionados. Sin embargo, cada uno de estos valores básicos constituye un prototipo local, en torno al cual se construye un complejo de matices semánticos derivados del prototipo. La idea final es que los hablantes son conscientes de los cuatro postulados valores básicos, pero que también distinguen matices más detallados, como son las ideas de 'causa', 'agente', 'instrumento', 'finalidad', 'cualidad', etc. Es decir, "de" constituye un elemento polisémico complejo cuya estructura semántica puede describirse como una semejanza de familia centrada en cuatro valores básicos en torno a los cuales se encuentra una serie de matices más específicos, que también constituyen valores propios de la preposición. Creemos, además, que esta caracterización semántica es válida para todas las épocas de la historia del español, con unas pequeñas modificaciones en el peso relativo de los distintos matices, lo cual está relacionado con la observada variación diacrónica en el uso de "de".

Genetic transformation of Brassica nigra by agrobacterium based vector and direct plasmid uptake

Genetic transformation systems have been established for Brassica nigra (cv. IC 257) by using an Agrobacterium binary vector as well as by direct DNA uptake of a plasmid vector. Both the type of vectors carried nptII gene and gus gene. For Agrobacterium mediated transformation, hypocotyl tissue explants were used, and up to 33% of the explants produced calli on selection medium. All of these expressed B-glucuronidase gene on histochemical staining. Protoplasts isolated from hypocotyl tissues of seedlings could be transformed with a plasmid vector by FEG mediated uptake of vector DNA. A number of fertile kanamycin resistant plants were obtained using both the methods, and their transformed nature was confirmed by Southern blot analysis and histochemical staining for GUS. Backcrossed and selfed progenies of these transformed plants showed the presence of npt and gus genes.

Absorption, translocation and metabolism of fluchloralin in groundnut (Arachis hypogaea) and pigweed (Amaranthus viridis)

Root absorption and translocation of [C-14]fluchloralin were determined in groundnut (Arachis hypogaea L.) cv. TMV-2 and pigweed (Amaranthus viridis L.) grown in nutrient solution culture under greenhouse conditions. Root-applied fluchloralin toxicity to groundnut and pigweed was also examined. A growth reduction of 50% occurred in groundnut that received fluchloralin at a concentration of 9.0 mum. Root absorption was similar for both groundnut and pigweed at one day after application (DAA), but groundnut absorbed about twice the amount of fluchloralin during 4 and 8 days of continuous application, compared with pigweed. Groundnut absorbed 25% of the total applied fluchloralin after 8 days. Translocation to leaves from treated roots was low and roots of groundnut contained 80% of the total absorbed C-14, 8 DAA. Contrary to the observations in groundnut, transport from the roots and leaves following root application in pigweed was rapid: 1 and 8 DAA, leaves of pigweed contained 45 and 70% of the total absorbed C-14, respectively. Different patterns of fluchloralin metabolism were observed in pigweed and groundnut. Pigweed metabolized most of the fluchloralin absorbed by roots. The fluchloralin tolerance of pigweed could partially be accounted for by absorption, translocation and metabolism.

Chloroplastic glutamine synthetase from normal and water stressed safflower (Carthamus tinctorius L.) leaves

The chloroplastic isoform of glutamine synthetase (GS(2), EC 6.3.1.2) from normal and water stressed safflower (Carthamus tinctorius L. cv.A-300) leaves has been purified to apparent electrophoretic homogeneity by a procedure involving anion-exchange, hydrophobic and size-exclusion chromatography followed by electroelution of the protein from preparative polyacrylamide gels. The observed molecular weight of the native protein varied from 305-330 kDa depending on the sizing column employed. The native protein is composed of 44 kDa subunits. Under conditions of saturating ammonium and at ATP levels of 0.1-10 mM, double-reciprocal plots with respect to glutamate are biphasic and concave downward at high concentrations of the varied substrate for normal enzyme but are linear for enzyme from water-stressed plants. Under subsaturating ATP levels, K-Glu is over 18-fold lower for enzyme from stressed leaves. The K-m, (ATP) varies with Mg2+ levels in the assay mixture. Double-reciprocal plots of initial velocity with respect to ATP at changing fixed levels of NH4+ are linear for normal enzyme but are curved upwards for enzyme from stressed leaves. Initial velocity data of 1/v vs. 1/ammonium for the enzyme from both the sources are non-linear (curved upwards) when ATP is saturating. At subsaturating ATP levels, the data are linear for normal enzyme but are still non-linear for the enzyme from stressed leaves. The results obtained suggest positively cooperative binding of NH4+ A V-max(/2) value of 3.6 mM for Mg2+ was obtained at 5 mM ATP. The isoelectric point of the native protein from normal and stressed leaves was determined to be, respectively, 5.6 and 6.1. The mixed competitive and competitive inhibitors, methionine sulfoximine and ADP and K-i values of 0.086 mM (0.017 for the enzyme from stressed leaves) and 2.15 mM (1.70 for the enzyme from stressed leaves), respectively. Enzyme from stressed leaves is not inhibited by 5 mM proline. The observed kinetic constants of GS(2) from normal and water stressed safflower seedlings are discussed in relation to the known water-stress tolerance of this crop plant.

First total synthesis of (±)-myltayl-4(12)-ene and single-crystal X-ray structure of exo-12-normyltaylan-4-yl 4-nitrobenzoate

The total synthesis of the unusual sesquiterpene (+/-)-myltayl-4(12)-ene 3 starting from the readily available cyclogeraniol 5 and the single-crystal X-ray structure of the 4-nitrobenzoate 12 of the noralcohol 11 are described.

A note on the choice of interfacial friction angle

Estimates of interfacial friction angle (delta) are necessary for the design of retaining structures and deep foundations, Recommendations in the literature regarding delta values are often contradictory and are therefore not easy to apply in geotechnical design, A critical examination of past studies in terms of data generation techniques used and conclusions drawn indicates that two distinctly different test procedures/techniques have been evolved. The interfacial situation in practice can also be categorized into two broad types, These two types of interface problems in geotechnical engineering are (a) the structure is placed on the free surface of prepared fill (type A situation) and (b) the fill is placed against the material surface which functions as a confined boundary (type B situation), The friction angle delta depends on the surface roughness of the construction material, But in the type A situation, it is independent of density and its limiting maximum value (delta(lim)) is the critical state friction angle phi(cv). In the type B situation, it is dependent on density of the fill and its limiting maximum value is the peak angle of internal friction phi(p) of the fill.

Expression of the Japanese encephalitis virus NS3 and NS2b proteins as glutathione S-transferase fusions

Flaviviruses generate their structural and nonstructural proteins by proteolytic processing of a single large polyprotein precursor. These proteolytic events are brought about both by host cell signalase and a virally encoded protease. The virally encoded proteolytic activity has been shown to reside within the nonstructural protein 3 (NS3) and requires the product of the nonstructural 2b (NS2b) gene. In order to obtain sufficient quantities of pure NS2b and NS3 proteins for kinetic analysis, we have expressed both these proteins in recombinant systems as fusions to glutathione S-transferase (GST). The fusion constructs were driven by the strong bacteriophage T7 promoter. Transfection of these constructs into the African green monkey kidney cell line CV-1 previously infected with a recombinant vaccinia virus expressing the T7 RNA polymerase resulted in synthesis of the fusion proteins. Both the fusion proteins could be purified to homogeneity in a single step using a glutathione agarose affinity matrix.

ChemInform Abstract: Simple and Regioselective Reductive Cleavage of Tetrahydropyranyl Ethers to Alcohols

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.