Questa ricerca è stata finanziata dal Fondo di Scienze austriaca (FWF): P23681-B16.

1. Preparazione di materiali vegetali <ol><li> Utilizzare sia foglie o piante intere di materiale vegetale soggetto (Hosta spp. O Phaseolus vulgaris). </li></ol> 2. Preparazione di Water Solutions contenenti ghiaccio nucleazione attivi (INA) Batteri <ol><li> Culture batterio INA, Pseudomonas syringae (Strain Cit-7) in capsule di Petri a 25 ° C su Pseudomonas Agar F preparato con 10 g / L di 100% glicerolo per la direzione del produttore. </li><li> Dopo culture sono cresciuti sufficientemente, posto a 4 ° C fino al momento dell&#39;uso, ma mantenere a 4 ° C per due giorni prima di garantire un elevato livello di attività nucleazione del ghiaccio. </li><li> Raschiare batteri da una singola placca dalla superficie del agar con una plastica, monouso o riutilizzabile spatola metallica al momento dell&#39;uso e posto in 10-15 ml di acqua deionizzata in un 25 ml cuvetta monouso. La concentrazione deve essere nel range di 1 x 07-01 Ottobre x 10 9 · </sup&gt; Ml -1. La soluzione apparirà nuvoloso. Non è necessario per confermare la concentrazione utilizzando un emocitometro o spettrofotometro, la concentrazione deve essere solo approssimativa. </li><li> Agitare la cuvetta per un minimo di 10 secondi per distribuire i batteri. Nota: la concentrazione specifica della miscela INA risultante non è importante e il protocollo descritto fornirà più di un adeguato livello di attività di nucleazione del ghiaccio. Questa miscela di batteri INA e acqua sarà utilizzato in seguito negli esperimenti di nucleazione. </li></ol> 3. Impostazione di un esperimento di congelamento <ol><li> Posizionare la telecamera a infrarossi alta risoluzione (SC-660) all&#39;interno della scatola acrilica di protezione in modo che i progetti lente attraverso l&#39;apertura nella parte anteriore della scatola e sui cavi la fotocamera a un dispositivo di uscita laptop o la registrazione attraverso l&#39;apertura posteriore della scatola . Fissare il coperchio della scatola e posizionare la scatola dentro la camera ambientale o congelatore in una posizione che saranno tuttiow il materiale vegetale soggetto di essere visto. <ol><li> Fornire un fondo scuro intorno al materiale vegetale da lungo le pareti della camera con cartoncino nero per evitare interferenze da energia infrarossa riflessa. </li><li> Montare la camera con illuminazione a LED per ridurre al minimo il riscaldamento dalla fonte di luce quando è richiesta la registrazione di immagini in lunghezze d&#39;onda visibili. Solo un minimo di illuminazione, ad esempio una luce di armadio a batteria o altro piccolo dispositivo LED, è necessaria per gli impianti siano visibili dalla telecamera. <ol><li> Una volta prese le immagini visibili del materiale vegetale soggetto, spegnere l&#39;illuminazione a LED. Distribuire tutte le connessioni esterne cablate (collegamento firewire al calcolatore, cavo di alimentazione, ecc) alla fotocamera tramite un porto o altra apertura nella camera. </li></ol></li><li> Riempite ogni spazio nel porto o apertura con materiale isolante in schiuma per evitare o ridurre i gradienti di temperatura all&#39;interno della camera. Impostare la temperatura iniziale della camera a 1 ° C. </li></ol></li><li> Allineate piante o parti di piante in modo che il materiale vegetale è nel campo di vista-della telecamera e il materiale vegetale è visibile sulla schermata di visualizzazione remota o all&#39;interno del software scelto. </li><li> Consentire piante equilibrare a 1 ° C per 30 minuti a 1 ora, a seconda delle dimensioni del materiale vegetale, prima di iniziare un esperimento di congelamento controllato. Questo assicura che la temperatura della pianta non ritardo temperatura dell&#39;aria da molti gradi dopo l&#39;inizio dell&#39;esperimento di congelamento. Equilibrazione viene raggiunta quando la temperatura del materiale vegetale è entro 0,5 ° C di temperatura dell&#39;aria. <ol><li> Posizionare uno strato di isolamento Styrofoam sulla parte superiore del terreno di piante in vaso se vengono utilizzate piante in vaso. Una volta che le piante sono equilibrati, iniziare il raffreddamento della camera. Nota: Lo strato di isolante sulla superficie del terreno della pentola riduce la quantità di continua perdita di calore dal piatto per l&#39;aria circostante l&#39;impianto, e previene le radici da freezing, in quanto ciò non si verificano in genere durante un evento gelo in natura a causa della massiccia serbatoio di calore residuo presente nel terreno. </li></ol></li><li> Impostare i parametri della telecamera desiderati (tavolozza dei colori, range di temperatura, specifiche aree di interesse, ecc), come discusso in 3.4.1-3.4.4. <ol><li> Selezionare la tavolozza arcobaleno per visualizzare le variazioni di temperatura durante la visualizzazione dell&#39;immagine live. </li><li> Impostare l&#39;intervallo di temperatura di 5 ° C regolando la barra di temperatura posta appena sotto l&#39;immagine nel software. </li><li> Scegliere la scala lineare (algoritmo) per convertire i dati infrarossi nell&#39;immagine a falsi colori come definito dalla palette selezionata (arcobaleno) e impostare l&#39;intervallo di temperature di 5 ° C e per monitorare automaticamente in base all&#39;immagine. In alternativa, regolare la gamma insieme manualmente durante la conduzione dell&#39;esperimento. <ol><li> Utilizzare la temperatura di un punto specifico o la temperatura media all&#39;interno dell&#39;area definita di interesse fornito dalla softwsono. Recuperare i dati di temperatura di tutti i pixel dalla sequenza video registrato o da informazioni incorporate nel file di immagine. La figura 3 mostra una tipica schermata dal software ResearchIR. </li></ol></li><li> Posizionare un cursore su una posizione sul tessuto vegetale che rappresenta un punto di interesse specifico. Definire l&#39;area di interesse come punti (1 -3 pixel), scatole, linee, ellissi o cerchi. Molteplici combinazioni di punti e forme possono essere posizionati sopra l&#39;immagine. </li></ol></li><li> Registrazione di una sequenza video <ol><li> Impostare la fotocamera per registrare a 60 Hz e per la registrazione di essere fermato manualmente. </li><li> Indicare la posizione del computer o disco esterno dove verrà posizionato il file video registrato. </li><li> Iniziare la registrazione. Nota: La registrazione di un disco rigido esterno altamente è raccomandata in quanto verranno generati file video di grandi dimensioni. I file video registrati possono essere successivamente modificati per includere solo la parte contenente il neinformazioni sario. Questo ridurrà di molto la dimensione del file. </li><li> Abbassare la temperatura della camera incrementale di 0,5 -1.0 ° C. Attendere che la temperatura dell&#39;impianto equilibra con la temperatura dell&#39;aria e abbassare di nuovo la temperatura di 0,5-1,0 ° C. A seconda della massa del tessuto vegetale viene osservato e la sua morfologia, equilibramento può richiedere da 10 a 15 min. Così, dando una velocità di raffreddamento di circa 4 ° C / hr. </li><li> Continuare in questo modo fino a quando l&#39;impianto si blocca e le osservazioni sono stati completati. Terminare la registrazione quando il processo di congelamento è stato completato. Nota: Il tessuto vegetale è equilibrata con temperatura dell&#39;aria quando il materiale vegetale e sfondo sono dello stesso colore poiché sono alla stessa temperatura. Poiché la temperatura del fondo e la temperatura del tessuto vegetale sono gli stessi, può essere difficile visualizzare il materiale vegetale fino a nuovo abbassare la temperatura e c&#39;è differenza di temperatura tra il tessuto vegetale etemperatura ir. </li></ol></li></ol>

<ol>
	<li>Taschler, D., Beikircher, B., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Frost+resistance+and+ice+nucleation+in+leaves+of+five+woody+timberline+species+measured+in+situ+during+shoot+expansion.">Frost resistance and ice nucleation in leaves of five woody timberline species measured in situ during shoot expansion.</a> Tree Physiol. 24, 331-337 (2004).</li><li>Neuner, G., Hacker, J., L&#252;tz, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+formation+and+propagation+in+alpine+plants.">Ice formation and propagation in alpine plants.</a> Plants in alpine regions: Cell Physiology of adaptation and survival strategies. , 163-174 (2012).</li><li>Ladinig, U., Hacker, J., Neuner, G., Wagner, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+endangered+is+sexual+reproduction+of+high-mountain+plants+by+summer+frosts?+-+Frost+resistance,+frequency+of+frost+events+and+risk+assessment.">How endangered is sexual reproduction of high-mountain plants by summer frosts? - Frost resistance, frequency of frost events and risk assessment.</a> Oecologia. 171, 743-760 (2013).</li><li>Wisniewski, M. E., Gusta, L. V., Fuller, M. P., Karlson, D., Gusta, L. V., Wisniewski, M. E., Tanino, K. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+nucleation,+propagation+and+deep+supercooling:+the+lost+tribes+of+freezing+studies.">Ice nucleation, propagation and deep supercooling: the lost tribes of freezing studies.</a> Plant Cold Hardiness: from the laboratory to the field. , 1-11 (2009).</li><li>Bokhurst, S., Bjerke, J. W., Davey, M. P., Taulavuori, K., Taulavuori, E., Laine, K., Callaghan, T. V., Phoenix, G. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impacts+of+extreme+winter+warming+events+on+plant+physiology+in+a+sub-Arctic+heath+community.">Impacts of extreme winter warming events on plant physiology in a sub-Arctic heath community.</a> Physiol. Plant. 140, 128-140 (2010).</li><li>Taulavuori, K., Laine, K., Taulavuori, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+studies+on+Vaccinium+myrtillus.and+Vaccinium+vits-idea.in+relation+to+air+pollution+and+global+change+at+northern+high+latitudes:+A+review.">Experimental studies on Vaccinium myrtillus.and Vaccinium vits-idea.in relation to air pollution and global change at northern high latitudes: A review.</a> Env. Exp. Bot. 87, 191-196 (2013).</li><li>Wisniewski, M., Glenn, D. M., Gusta, L., Fuller, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+infrared+thermography+to+study+freezing+in+plants.">Using infrared thermography to study freezing in plants.</a> HortScience. 43, 1648-1651 (2008).</li><li>Gu, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+2007+eastern+US+spring+freeze:+increased+cold+damage+in+a+warming+world.">The 2007 eastern US spring freeze: increased cold damage in a warming world.</a> BioScience. 58, 253-262 (2008).</li><li>Augspurger, C. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spring+warmth+and+frost:+phenology,+damage,+and+refoliation+in+a+temperate+deciduous+forest.">Spring warmth and frost: phenology, damage, and refoliation in a temperate deciduous forest.</a> Func. Ecol. 23, 1031-1039 (2007).</li><li>Neuner, G., Erler, A., Hacker, J., Ladinig, U., Wagner, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Frost+resistance+of+reproductive+tissues+in+various+reproductive+stages+of+high+alpine+plant+species.">Frost resistance of reproductive tissues in various reproductive stages of high alpine plant species.</a> Physiol. Plant. 147, 88-100 (2013).</li><li>Skre, O., Taulavuori, K., Taulavuori, E., Nilsne, J., Igeland, B., Laine, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+importance+of+hardening+and+winter+temperature+for+growth+in+mountain+birch+populations.">The importance of hardening and winter temperature for growth in mountain birch populations.</a> Env. Exp. Bot. 62, 254-266 (2008).</li><li>H&#228;nninen, H., Tanino, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tree+seasonality+in+a+warming+climate.">Tree seasonality in a warming climate.</a> Trends in Plant Science. 16, 412-416 (2011).</li><li>Katz, R. W., Brown, B. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extreme+events+in+a+changing+climate:+variability+is+more+important+than+averages.">Extreme events in a changing climate: variability is more important than averages.</a> Climate Change. 21, 289-302 (1992).</li><li>Wisniewski, M., Gusta, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Understanding+plant+cold+hardiness:+an+opinion.">Understanding plant cold hardiness: an opinion.</a> Physiol. Plant. 147, 4-14 (2013).</li><li>Wisniewski, M., Gusta, L., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adaptive+mechanisms+of+freeze+avoidance+in+plants.+A+brief+update.">Adaptive mechanisms of freeze avoidance in plants. A brief update.</a> Env. Exp. Bot. 99, 133-140 (2014).</li><li>Burke, M. J., Gusta, L. V., Quamme, H. A., Weiser, C. J., Li, P. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Freezing+and+injury+in+plants.">Freezing and injury in plants.</a> Annu. Rev. Plant Physiol. 27, 507-528 (1976).</li><li>Ishikawa, M., Price, W. S., Ide, H., Arata, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Visualization+of+freezing+behaviors+in+leaf+and+flower+buds+of+full-moon+maple+by+nuclear+magnetic+resonance+microscopy.">Visualization of freezing behaviors in leaf and flower buds of full-moon maple by nuclear magnetic resonance microscopy.</a> Plant Physiol. 115 (4), 1515-1524 (1997).</li><li>Ishikawa, M., Sakai, A., Li, P. H., Sakai, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characteristics+of+freezing+avoidance+in+comparison+with+freezing+tolerance:+a+demonstration+of+extra-organ+freezing.">Characteristics of freezing avoidance in comparison with freezing tolerance: a demonstration of extra-organ freezing.</a> Plant cold hardiness and freezing stress. , 325-340 (1982).</li><li>Buchner, O., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Freezing+cytorrhysis+and+critical+temperature+thresholds+for+photosystem+II+in+the+peat+moss+Sphagnum+capillifolium.">Freezing cytorrhysis and critical temperature thresholds for photosystem II in the peat moss Sphagnum capillifolium.</a> Protoplasma. 243 (1), 63-71 (2010).</li><li>Pearce, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+ice+and+cell+shape+in+frost-stressed+cereal+leaves:+A+low+temperature+scanning+electron+microscopy+study.">Extracellular ice and cell shape in frost-stressed cereal leaves: A low temperature scanning electron microscopy study.</a> Planta. 175, 313-324 (1988).</li><li>Ashworth, E. N., Anderson, J. A., Davis, G. A., Lightner, G. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+formation+in+Prunus+persica.+under+field+conditions.">Ice formation in Prunus persica. under field conditions.</a> J. Am. Soc. Hort. Sci. 110 (3), 322-324 (1985).</li><li>Ashworth, E. N., Davis, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+formation+in+woody+plants+under+field+conditions.">Ice formation in woody plants under field conditions.</a> HortSci. 21, 1233-1234 (1986).</li><li>Pramsohler, M., Hacker, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Freezing+pattern+and+frost+killing+temperature+of+apple+(Malus+domestica.)+wood+under+controlled+conditions+and+in+nature.">Freezing pattern and frost killing temperature of apple (Malus domestica.) wood under controlled conditions and in nature.</a> Tree Physiol. 32 (7), 819-828 (2012).</li><li>Wisniewski, M., Lindow, S. E., Ashworth, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Observations+of+ice+nucleation+and+propagation+in+plants+using+infrared+video+thermography.">Observations of ice nucleation and propagation in plants using infrared video thermography.</a> Plant Physiol. 113 (2), 327-334 (1997).</li><li>Lutze, J. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elevated+atmospheric+[CO2]+promotes+frost+damage+in+evergreen+tree+seedlings.">Elevated atmospheric [CO2] promotes frost damage in evergreen tree seedlings.</a> Plant Cell Environ. (6), 631-635 (1998).</li><li>Ball, M. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Space+and+time+dependence+of+temperature+and+freezing+in+evergreen+leaves).">Space and time dependence of temperature and freezing in evergreen leaves).</a> Func. Plant Biol. 29 (11), 1259-1272 (2002).</li><li>Sekozawa, Y., Sugaya, S., Gemma, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Observations+of+ice+nucleation+and+propagation+in+flowers+of+Japanese+Pear+(Pyrus+Pyrifolia).">Observations of ice nucleation and propagation in flowers of Japanese Pear (Pyrus Pyrifolia).</a> Nakai) using infrared video. 73 (1), 1-6 (2004).</li><li>Hacker, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+propagation+in+plants+visualized+at+the+tissue+level+by+IDTA+(infrared+differential+thermal+analysis).">Ice propagation in plants visualized at the tissue level by IDTA (infrared differential thermal analysis).</a> Tree Physiol. 27, 1661-1670 (2007).</li><li>Hacker, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+propagation+in+dehardened+alpine+plant+species+studied+by+infrared+differential+thermal+analysis+(IDTA).">Ice propagation in dehardened alpine plant species studied by infrared differential thermal analysis (IDTA).</a> Arc. Antarc. Alp. Res. 40 (4), 660-670 (2008).</li><li>Hacker, J., Spindelb&#246;ck, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mesophyll+freezing+and+effects+of+freeze+dehydration+visualized+by+simultaneous+measurement+of+IDTA+and+differential+imaging+chlorophyll+fluorescence.">Mesophyll freezing and effects of freeze dehydration visualized by simultaneous measurement of IDTA and differential imaging chlorophyll fluorescence.</a> Plant Cell Environ. 31, 1725-1733 (2008).</li><li>Neuner, G., XU, B., Hacker, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Velocity+and+pattern+of+ice+propagation+and+deep+supercooling+in+woody+stems+of+Castanea+sativa.,+Morus+nigra.+and+Quercus+robur.+measured+by+IDTA.">Velocity and pattern of ice propagation and deep supercooling in woody stems of Castanea sativa., Morus nigra. and Quercus robur. measured by IDTA.</a> Tree Physiol. 30, 1037-1045 (2010).</li><li>Hacker, J., Ladinig, U., Wagner, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflorescences+of+alpine+cushion+plants+freeze+autonomously+and+may+survive+subzero+temperatures+by+supercooling.">Inflorescences of alpine cushion plants freeze autonomously and may survive subzero temperatures by supercooling.</a> Plant Sci. 180, 149-156 (2011).</li><li>Kuprian, E., Briceno, V., Wagner, J., Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ice+barriers+promote+supercooling+and+prevent+frost+injury+in+reproductive+buds,+flowers+and+fruits+of+alpine+dwarf+shrubs+throughout+the+summer.">Ice barriers promote supercooling and prevent frost injury in reproductive buds, flowers and fruits of alpine dwarf shrubs throughout the summer.</a> Env. Exp. Bot. 106, 4-12 (2014).</li><li>Wisniewski, M., Glenn, D. M., Fuller, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+a+hydrophobic+particle+film+as+a+barrier+to+extrinsic+ice+nucleation+in+tomato+plants.">Use of a hydrophobic particle film as a barrier to extrinsic ice nucleation in tomato plants.</a> HortScience. 127, 358-364 (2002).</li><li>Aryal, B., &amp;Neuner, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Leaf+wettability+decreases+along+an+extreme+altitudinal+gradient.">Leaf wettability decreases along an extreme altitudinal gradient.</a> Oecologia. 162, 1-9 (2010).</li><li>Wisniewski, M., Fuller, M. P., Margesin, R., Schinner, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Ice nucleation and deep supercooling in plants: new insights using infrared thermography. In: Cold adapted organisms. Ecology, physiology, enzymologyandmolecularbiology. , 105-118 (1999).</li><li>Franks, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Biophysics and biochemistry at low temperatures. , (1985).</li><li>Neuner, G., Kuprian, E., Hincha, D., Zuther, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Infrared+thermal+analysis+of+plant+freezing+processes.">Infrared thermal analysis of plant freezing processes.</a> Methods in Molecular Biology: Plant Cold Acclimation. , 91-98 (2014).</li></ol>

Gli autori non hanno interessi finanziari in competizione o conflitti di interesse.

L&#39;acqua ha la capacità di Supercool a temperature inferiori a 0 ° C e la temperatura alla quale l&#39;acqua si congela può essere molto variabile. 36 Il limite di temperatura per sottoraffreddamento di acqua pura è di circa -40 ° C ed è definito come il punto di nucleazione omogenea. Quando l&#39;acqua gela A temperature superiori a -40 ° C si è portato dalla presenza di eterogeneo nucleatori che consentono piccoli embrioni di ghiaccio di forma che poi servire come catalizzatore per la formazione ...

Temperature di congelamento che si verificano quando le piante sono in attiva crescita può essere letale, in particolare se la pianta ha poca o nessuna tolleranza congelamento. Tali eventi gelo spesso hanno effetti devastanti sulla produzione agricola e possono svolgere un ruolo importante nel plasmare la struttura della comunità in popolazioni naturali di piante, in particolare in montagna, gli ecosistemi sub-artiche e artiche 1-6. Gli episodi di gravi gelate primaverili hanno avuto maggiori impatti sulla produzione di frutta negli Stati Uniti e in Sud America, negli ultimi anni 7-9 e sono stati esacerbati dalla precoce inizio della stagione calda seguita da più tipici basse temperature medie. Il clima caldo precoce induce i germogli di rompere, attivando la crescita di nuovi germogli, foglie e fiori che hanno tutti molto poco o nessun tolleranza al freddo 1,3,10-12. Tali condizioni meteorologiche imprevedibili sono stati segnalati per essere un riflesso diretto del cambiamento climatico in corso e si prevede che essere un modello di tempo comune per Foresfuturo eeable 13. Gli sforzi per fornire tecniche di gestione economiche, efficaci e rispettosi dell&#39;ambiente o prodotti agrochimici in grado di fornire una maggiore tolleranza al freddo hanno avuto un successo limitato per una serie di motivi, ma questo può essere in parte attribuito alla natura complessa di congelare la tolleranza e il congelamento dei meccanismi di evitamento nelle piante. 14 I meccanismi di adattamento associata a sopravvivenza gelo nelle piante sono stati tradizionalmente divisi in due categorie, il congelamento la tolleranza e il congelamento di evitamento. La prima categoria è associata con meccanismi biochimici regolati da uno specifico set di geni che permettono alle piante di tollerare tensioni associate con la presenza e l&#39;effetto dehydrative di ghiaccio nei suoi tessuti. Mentre quest&#39;ultima categoria è tipicamente, ma non esclusivamente, associato ad aspetti strutturali di una pianta che determinano se, quando, e dove si forma il ghiaccio in un impianto di 14. Nonostante la prevalenza di evasione freeze come un annuncioMeccanismo aptive, poca ricerca è stata dedicata negli ultimi tempi per la comprensione dei meccanismi e regolazione di evasione freeze sottostanti. Il lettore si riferisce a una recente revisione 15 per maggiori dettagli in merito. Mentre la formazione di ghiaccio a basse temperature può sembrare un processo semplice, molti fattori contribuiscono a determinare la temperatura alla quale il ghiaccio nucleates in tessuti vegetali e come si diffonde all&#39;interno della pianta. Parametri come la presenza di ghiaccio estrinseca e intrinseca nucleatori, eterogenei vs eventi nucleazione omogenea, termico-isteresi (antigelo) proteine, la presenza di zuccheri specifici e altri osmoliti, e una serie di aspetti strutturali della pianta possono tutti giocare un significativo ruolo nel processo di congelamento in impianti. Collettivamente, questi parametri influenzano la temperatura alla quale una pianta gela, in cui viene iniziata ghiaccio e come cresce. Essi possono anche influenzare la morfologia dei cristalli di ghiaccio risultanti.Vari metodi sono stati utilizzati per studiare il processo di congelamento in impianti in condizioni di laboratorio, compresa la spettroscopia di risonanza magnetica nucleare (NMR) 16, la risonanza magnetica (MRI) 17, crio-microscopia 18-19, e bassa temperatura di microscopia elettronica a scansione (LTSEM ). 20 Congelamento di piante intere in laboratorio e sul campo, però, è soprattutto stata monitorata con termocoppie. L&#39;uso di termocoppie per studiare congelamento si basa sulla liberazione di calore (entalpia di fusione) quando l&#39;acqua subisce una transizione di fase da liquido a solido. Congelamento viene quindi registrato come un evento esotermica. 21-23 Sebbene termocoppie sono il tipico metodo di scelta nello studio congelamento nelle piante, il loro uso ha molte limitazioni che limitano la quantità di informazioni ottenute durante un evento di congelamento. Ad esempio, con termocoppie è difficile quasi impossibile determinare dove viene iniziata ghiaccio in piante, come si propaga,se si propaga ad una frequenza costante, e se alcuni tessuti rimangono liberi di ghiaccio. I progressi in alta risoluzione termografia a infrarossi (HRIT) 24-27, però, hanno aumentato significativamente la capacità di ottenere informazioni sul processo di congelamento di piante intere, specialmente se usata in modalità di imaging differenziale. 28-33 Nella presente relazione, abbiamo descrivere l&#39;uso di questa tecnologia per studiare vari aspetti del processo di congelamento e vari parametri che influiscono dove e ghiaccio ea quale temperatura è iniziata nelle piante. Un protocollo sarà presentato che dimostrare la capacità del batterio di ghiaccio-nucleazione-attiva (INA), Pseudomonas syringae (Cit-7) per agire come un nucleator estrinseca avvio congelamento in una pianta erbacea ad una temperatura elevata sotto zero. Ad alta risoluzione telecamera a infrarossi Il protocollo e gli esempi documentati in questo rapporto utilizzano una alta risoluzione infrarossiVideo radiometro. Il radiometro (figura 1) fornisce un insieme di immagini spettro all&#39;infrarosso e nel visibile e dati di temperatura. La risposta spettrale della fotocamera è nel range di 7,5-13,5 pm e fornisce 640 x 480 pixel. Immagini spettro visibile generati dalla fotocamera incorporata può essere fuso con IR-immagini in tempo reale, che facilita l&#39;interpretazione dei complessi, immagini termiche. Una gamma di lenti per la macchina fotografica può essere usato per fare close-up e osservazioni microscopiche. La telecamera può essere utilizzata in una modalità autonoma, o interfacciato e controllato con un computer portatile utilizzando il software propietari. Il software può essere utilizzato per ottenere una varietà di dati termici incorporati nei video registrati. E &#39;importante notare che una grande varietà di radiometri infrarossi sono disponibili in commercio. Pertanto, è essenziale che il ricercatore discutere l&#39;applicazione prevista con un tecnico esperto del prodotto e che il ricercatore testare la capacità di qualsiasi specific radiometro a fornire le informazioni necessarie. Il radiometro di imaging utilizzata nel protocollo descritto è posto in una scatola di acrilico (Figura 2) isolato con Styrofoam i n fine di dissuadere l&#39;esposizione alla condensa durante i protocolli di riscaldamento e raffreddamento. Questa protezione non è necessaria per tutte le telecamere o le applicazioni.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Infrared Camera</td>
			<td>FLIR</td>
			<td>SC-660</td>
			<td>Many models available depending on application</td>
		</tr>
		<tr>
			<td>Infrared Analytical Software</td>
			<td>FLIR</td>
			<td>ResearchIR 4.10.2.5</td>
			<td align="right">$3,500</td>
		</tr>
		<tr>
			<td rowspan="2">Pseudomonas syringae (strain Cit-7)</td>
			<td rowspan="2"></td>
			<td rowspan="2"></td>
			<td rowspan="2"><a href="mailto:icelab@berkeley.edu">Kindly provided by Dr. Steven Lindow, University of California&#160; Berkeley icelab@berkeley.edu</a></td>
		</tr>
		<tr>
		</tr>
		<tr>
			<td>Pseudomonas Agar F</td>
			<td>Fisher Scientific</td>
			<td>DF0448-17-1</td>
			<td></td>
		</tr>
	</tbody>
</table>

the use of high-resolution infrared thermography (hrit) for the study of ice nucleation and ice propagation in plants

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events often have devastating effects on agricultural production and can also play an important role in shaping community structure in natural populations of plants, especially in alpine, sub-arctic, and arctic ecosystems. Therefore, a better understanding of the freezing process in plants can play an important role in the development of methods of frost protection and understanding mechanisms of freeze avoidance. Here, we describe a protocol to visualize the freezing process in plants using high-resolution infrared thermography (HRIT). The use of this technology allows one to determine the primary sites of ice formation in plants, how ice propagates, and the presence of ice barriers. Furthermore, it allows one to examine the role of extrinsic and intrinsic nucleators in determining the temperature at which plants freeze and evaluate the ability of various compounds to either affect the freezing process or increase freezing tolerance. The use of HRIT allows one to visualize the many adaptations that have evolved in plants, which directly or indirectly impact the freezing process and ultimately enables plants to survive frost events.

Attività di Ice-nucleazione del ghiaccio + batterio, Pseudomonas syringae (ceppo Cit-7) Una goccia 10 ml di acqua e 10 ml di acqua contenente P. syringae (Cit-7) sono stati collocati sulla superficie di una foglia abassiale Hosta (Hosta spp.) (Figura 4). Come illustrato, la goccia d&#39;acqua contenente batteri INA congelato prima ed era responsabile per indurre la foglia di congelare mentre la goccia d&#39;acqua sulla superficie del foglio è rim...

Watch this Scientific Journal Video about The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants at JoVE.com

The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

L&#39;uso di ad alta risoluzione termografia a infrarossi (HRIT) per lo studio di ghiaccio nucleazione del ghiaccio e propagazione nelle piante

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events ...

the-use-high-resolution-infrared-thermography-hrit-for-study-ice

Research

JoVE Journal

Environment

The Use of High-resolution Infrared Thermography (HRIT) for the Study of Ice Nucleation and Ice Propagation in Plants

9.5K Views.  Agricultural Research Service (USDA-ARS), Kearneysville, WV. Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

Video: The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

Cellulose degrading enzymes, cellulases, are targets of both research and industrial interests. The preponderance of these enzymes in difficult-to-culture organisms, such as hyphae-building fungi and anaerobic bacteria, has hastened the use of recombinant technologies in this field. Plant expression methods are a desirable system for large-scale production of enzymes and other industrially useful proteins. Herein, methods for the transient expression of a fungal endoglucanase, Trichoderma reesei Cel5A, in Nicotiana tabacum are demonstrated. Successful protein expression is shown, monitored by fluorescence using an mCherry-enzyme fusion protein. Additionally, a set of basic tests are used to examine the activity of transiently expressed T. reesei Cel5A, including SDS-PAGE, Western blotting, zymography, as well as&#160;fluorescence and dye-based substrate degradation assays. The system described here can be used to produce an active cellulase in a short time period, so as to assess the potential for further production in plants through constitutive or inducible expression systems.

Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

Tobacco plants were used to produce a fungal cellulase, TrCel5A, via a transient expression system. The expression could be monitored using a fluorescent fusion protein, and the protein activity was characterized post-expression.

Espressione ricombinante Cellulase Cel5A da<em&gt; Trichoderma reesei</em&gt; In piante di tabacco

Cellulose degrading enzymes, cellulases, are targets of both research and industrial interests. The preponderance of these enzymes in difficult-to-culture ...

Ricerca

Ambiente

Enhancement of the Initial Growth Rate of Agricultural Plants by Using Static Magnetic Fields

Electronic devices and high-voltage wires induce magnetic fields. A magnetic field of 1,300-2,500 Gauss (0.2 Tesla) was applied to Petri dishes containing seeds of Garden Balsam (Impatiens balsamina), Mizuna (Brassica rapa var. japonica), Komatsuna (Brassica rapa var. perviridis), and Mescluns (Lepidium sativum). We applied magnets under the culture dish. During the 4 days of application, we observed that the stem and root length increased. The group subjected to magnetic field treatment (n = 10) showed a 1.4 times faster rate of growth compared with the control group (n = 11) in a total of 8 days (p &#60;0.0005). This rate is 20% higher than that reported in previous studies. The tubulin complex lines did not have connecting points, but connecting points occur upon the application of magnets. This shows complete difference from the control, which means abnormal arrangements. However, the exact cause remains unclear. These results of growth enhancement of applying magnets suggest that it is possible to enhance the growth rate, increase productivity, or control the speed of germination of plants by applying static magnetic fields. Also, magnetic fields can cause physiological changes in plant cells and can induce growth. Therefore, stimulation with a magnetic field can have possible effects that are similar to those of chemical fertilizers, which means that the use of fertilizers can be avoided.

The goal of this protocol is to demonstrate the acceleration of the initial growth rate of plants by applying static magnetic fields with no external energy.

Valorizzazione del tasso di crescita iniziale delle Piante Agrarie mediante l&#39;utilizzo di campi magnetici statici

Electronic devices and high-voltage wires induce magnetic fields. A magnetic field of 1,300-2,500 Gauss (0.2 Tesla) was applied to Petri dishes containing ...

Use of a Battery of Chemical and Ecotoxicological Methods for the Assessment of the Efficacy of Wastewater Treatment Processes to Remove Estrogenic Potency

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in a watch list of environmental pollutants under the European Water Framework Directive. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. Much of the estrogenic potency of wastewater treatment plant (WWTP) effluents can be attributed to the discharge of steroid estrogens including estradiol (E2), EE2 and E1 due to incomplete removal of these substances at the treatment plant. An evaluation of the efficacy of wastewater treatment processes requires the quantitative determination of individual substances most often undertaken using chemical analysis methods. Most frequently used methods include Gas Chromatography-Mass Spectrometry (GCMS/MS) or Liquid Chromatography-Mass Spectrometry (LCMS/MS) using multiple reaction monitoring (MRM). Although very useful for regulatory purposes, targeted chemical analysis can only provide data on the compounds (and specific metabolites) monitored. Ecotoxicology methods additionally ensure that any by-products produced or unknown estrogenic compounds present are also assessed via measurement of their biological activity. A number of in vitro bioassays including the Yeast Estrogen Screen (YES) are available to measure the estrogenic activity of wastewater samples. Chemical analysis in conjunction with in vivo and in vitro bioassays provides a useful toolbox for assessment of the efficacy and suitability of wastewater treatment processes with respect to estrogenic endocrine disrupting compounds. This paper utilizes a battery of chemical and ecotoxicology tests to assess conventional, advanced and emerging wastewater treatment processes in laboratory and field studies.

Endocrine Disrupting Compounds (EDC) pose a substantial risk to the aquatic environment. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. The methodology provided in this paper allows for an assessment of the efficacy and suitability of wastewater treatment processes with respect to EDC removal.

Utilizzo di una batteria di chimici ed ecotossicologici metodi per la valutazione dell&#39;efficacia dei processi di trattamento delle acque reflue per rimuovere estrogenica Potenza

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in ...

Empirical, Metagenomic, and Computational Techniques Illuminate the Mechanisms by which Fungicides Compromise Bee Health

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered &#34;bee-safe,&#34; there is mounting evidence that fungicide residues in pollen are associated with bee declines (for both honey and bumble bee species). While the mechanisms remain relatively unknown, researchers have speculated that bee-microbe symbioses are involved. Microbes play a pivotal role in the preservation and/or processing of pollen, which serves as nutrition for larval bees. By altering the microbial community, it is likely that fungicides disrupt these microbe-mediated services, and thereby compromise bee health. This manuscript describes the protocols used to investigate the indirect mechanism(s) by which fungicides may be causing colony decline. Cage experiments exposing bees to fungicide-treated flowers have already provided the first evidence that fungicides cause profound colony losses in a native bumble bee (Bombus impatiens). Using field-relevant doses of fungicides, a series of experiments have been developed to provide a finer description of microbial community dynamics of fungicide-exposed pollen. Shifts in the structural composition of fungal and bacterial assemblages within the pollen microbiome are investigated by next-generation sequencing and metagenomic analysis. Experiments developed herein have been designed to provide a mechanistic understanding of how fungicides affect the microbiome of pollen-provisions. Ultimately, these findings should shed light on the indirect pathway through which fungicides may be causing colony declines.

Microbial consortia within bumble bee hives enrich and preserve pollen for bee larvae. Using next generation sequencing, along with laboratory and field-based experiments, this manuscript describes protocols used to test the hypothesis that fungicide residues alter the pollen microbiome, and colony demographics, ultimately leading to colony loss.

Empirica, metagenomica e tecniche computazionali illuminano i meccanismi attraverso i quali salute delle API di fungicidi compromesso

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered ...

Automated, High-resolution Mobile Collection System for the Nitrogen Isotopic Analysis of NOx

Nitrogen oxides (NOx = NO + NO2) are a family of atmospheric trace gases that have great impact on the environment. NOx concentrations directly influence the oxidizing capacity of the atmosphere through interactions with ozone and hydroxyl radicals. The main sink of NOx is the formation and deposition of nitric acid, a component of acid rain and a bioavailable nutrient. NOx is emitted from a mixture of natural and anthropogenic sources, which vary in space and time. The collocation of multiple sources and the short lifetime of NOx make it challenging to quantitatively constrain the influence of different emission sources and their impacts on the environment. Nitrogen isotopes of NOx have been suggested to vary amongst different sources, representing a potentially powerful tool to understand the sources and transport of NOx. However, previous methods of collecting atmospheric NOx integrate over long (week to month) time spans and are not validated for the efficient collection of NOx in relevant, diverse field conditions. We report on a new, highly efficient field-based system that collects atmospheric NOx for isotope analysis at a time resolution between 30 min and 2 hr. This method collects gaseous NOx in solution as nitrate with 100% efficiency under a variety of conditions. Protocols are presented for collecting air in urban settings under both stationary and mobile conditions. We detail the advantages and limitations of the method and demonstrate its application in the field. Data from several deployments are shown to 1) evaluate field-based collection efficiency by comparisons with in situ NOx concentration measurements, 2) test the stability of stored solutions before processing, 3) quantify in situ reproducibility in a variety of urban settings, and 4) demonstrate the range of N isotopes of NOx detected in ambient urban air and on heavily traveled roadways.

Automated, High-resolution Mobile Collection System for the Nitrogen Isotopic Analysis of NOx

Previous work suggests that the nitrogen isotopic composition of atmospheric nitrogen oxides might distinguish the influence of different sources in the environment. We report on an automated, mobile, field-based method for the high collection efficiency of atmospheric NOx for N isotopic analysis at an hourly time resolution.

Automatizzato, ad alta risoluzione mobile sistema di raccolta per l&#39;analisi isotopica di azoto di NO<sub&gt; x</sub

Nitrogen oxides (NOx = NO + NO2) are a family of atmospheric trace gases that have great impact on the environment. NOx concentrations directly influence ...

The Calibration and Use of Capacitance Sensors to Monitor Stem Water Content in Trees

Water transport and storage through the soil-plant-atmosphere continuum is critical to the terrestrial water cycle, and has become a major research focus area. Biomass capacitance plays an integral role in the avoidance of hydraulic impairment to transpiration. However, high temporal resolution measurements of dynamic changes in the hydraulic capacitance of large trees are rare. Here, we present procedures for the calibration and use of capacitance sensors, typically used to monitor soil water content, to measure the volumetric water content in trees in the field. Frequency domain reflectometry-style observations are sensitive to the density of the media being studied. Therefore, it is necessary to perform species-specific calibrations to convert from the sensor-reported values of dielectric permittivity to volumetric water content. Calibration is performed on a harvested branch or stem cut into segments that are dried or re-hydrated to produce a full range of water contents used to generate a best-fit regression with sensor observations. Sensors are inserted into calibration segments or installed in trees after pre-drilling holes to a tolerance fit using a fabricated template to ensure proper drill alignment. Special care is taken to ensure that sensor tines make good contact with the surrounding media, while allowing them to be inserted without excessive force. Volumetric water content dynamics observed via the presented methodology align with sap flow measurements recorded using thermal dissipation techniques and environmental forcing data. Biomass water content data can be used to observe the onset of water stress, drought response and recovery, and has the potential to be applied to the calibration and evaluation of new plant-level hydrodynamics models, as well as to the partitioning of remotely sensed moisture products into above- and belowground components.

The hydraulic capacitance of biomass is a key component of the vegetation water budget, which serves as a buffer against short and long-term drought stresses. Here, we present a protocol for the calibration and use of soil moisture capacitance sensors to monitor water content in the stems of large trees.

La calibrazione e l'uso di capacitanza sensori per monitorare il contenuto di acqua di staminali in alberi

Water transport and storage through the soil-plant-atmosphere continuum is critical to the terrestrial water cycle, and has become a major research focus ...

Stress Distribution During Cold Compression of Rocks and Mineral Aggregates Using Synchrotron-based X-Ray Diffraction

We report detailed procedures for performing compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus (D-DIA) coupled with synchrotron X-radiation. A cube-shaped sample assembly is prepared and compressed, at room temperature, by a set of four X-ray transparent sintered diamond anvils and two tungsten carbide anvils, in the lateral and the vertical planes, respectively. All six anvils are housed within a 250-ton hydraulic press and driven inward simultaneously by two wedged guide blocks. A horizontal energy dispersive X-ray beam is projected through and diffracted by the sample assembly. The beam is commonly in the mode of either white or monochromatic X-ray. In the case of white X-ray, the diffracted X-rays are detected by a solid-state detector array that collects the resulting energy dispersive diffraction pattern. In the case of monochromatic X-ray, the diffracted pattern is recorded using a two-dimensional (2-D) detector, such as an imaging plate or a charge-coupled device (CCD) detector. The 2-D diffraction patterns are analyzed to derive lattice spacings. The elastic strains of the sample are derived from the atomic lattice spacing within grains. The stress is then calculated using the predetermined elastic modulus and the elastic strain. Furthermore, the stress distribution in two-dimensions allow for understanding how stress is distributed in different orientations. In addition, a scintillator in the X-ray path yields a visible light image of the sample environment, which allows for the precise measurement of sample length changes during the experiment, yielding a direct measurement of volume strain on the sample. This type of experiment can quantify the stress distribution within geomaterials, which can ultimately shed light on the mechanism responsible for compaction. Such knowledge has the potential to significantly improve our understanding of key processes in rock mechanics, geotechnical engineering, mineral physics, and material science applications where compactive processes are important.

We report detailed procedures for compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus coupled with synchrotron X-radiation. Such experiments allow quantification of the stress distribution within samples, that ultimately sheds light on compaction processes in geomaterials.

Distribuzione delle sollecitazioni durante la compressione a freddo di rocce e inerti minerali mediante diffrazione di raggi x di sincrotrone-basato

We report detailed procedures for performing compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus (D-DIA) ...

Protocol for Producing Three-Dimensional Infrared Video of Freezing in Plants

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast make 2-dimensions (2D) infrared images somewhat difficult to interpret. Viewing an IR image or the video of plants freezing in 3 dimensions (3D) would allow a more accurate identification of sites for ice nucleation as well as the progression of freezing. In this paper, we demonstrate a relatively simple means to produce a 3D infrared video of a strawberry plant freezing. Strawberry is an economically important crop that is subjected to unexpected spring freeze events in many areas of the world. An accurate understanding of the freezing in strawberry will provide both breeders and growers with more economical ways to prevent any damage to plants during freezing conditions.
The technique involves a positioning of two IR cameras at slightly different angles to film the strawberry freezing. The two video streams will be precisely synchronized using a screen capture software that records both cameras simultaneously. The recordings will then be imported into the imaging software and processed using an anaglyph technique. Using red-blue glasses, the 3D video will make it easier to determine the precise site of ice nucleation on leaf surfaces.

Here, we present a protocol to image a strawberry plant freezing in 3 dimensions. Two infrared cameras positioned at slightly different angles are used to produce a red-blue anaglyph video to observe the freezing of the plant in 3 dimensions.

Protocollo per la produzione di Video tridimensionale a raggi infrarossi di congelamento in piante

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast ...

Long-term Video Tracking of Cohoused Aquatic Animals: A Case Study of the Daily Locomotor Activity of the Norway Lobster (Nephrops norvegicus)

We present a protocol related to a video-tracking technique based on the background subtraction and image thresholding that makes it possible to individually track cohoused animals. We tested the tracking routine with four cohoused Norway lobsters (Nephrops norvegicus) under light-darkness conditions for 5 days. The lobsters had been individually tagged. The experimental setup and the tracking techniques used are entirely based on the open source software. The comparison of the tracking output with a manual detection indicates that the lobsters were correctly detected 69% of the times. Among the correctly detected lobsters, their individual tags were correctly identified 89.5% of the times. Considering the frame rate used in the protocol and the movement rate of lobsters, the performance of the video tracking has a good quality, and the representative results support the validity of the protocol in producing valuable data for research needs (individual space occupancy or locomotor activity patterns). The protocol presented here can be easily customized and is, hence, transferable to other species where the individual tracking of specimens in a group can be valuable for answering research questions.

Long-term Video Tracking of Cohoused Aquatic Animals: A Case Study of the Daily Locomotor Activity of the Norway Lobster Nephrops norvegicus

Here we present a protocol to individually track animals over a long period of time. It uses computer vision methods to identify a set of manually constructed tags by using a group of lobsters as case study, simultaneously providing information on how to house, manipulate, and mark the lobsters.

A lungo termine dei Video rilevamento di Cohoused animali acquatici: un caso di studio dell'attività locomotrice quotidiana di scampi (Nephrops norvegicus)

We present a protocol related to a video-tracking technique based on the background subtraction and image thresholding that makes it possible to ...

Single-throughput Complementary High-resolution Analytical Techniques for Characterizing Complex Natural Organic Matter Mixtures

Natural organic matter (NOM) is composed of a highly complex mixture of thousands of organic compounds which, historically, proved difficult to characterize. However, to understand the thermodynamic and kinetic controls on greenhouse gas (carbon dioxide [CO2] and methane [CH4]) production resulting from the decomposition of NOM, a molecular-level characterization coupled with microbial proteome analyses is necessary. Further, climate and environmental changes are expected to perturb natural ecosystems, potentially upsetting complex interactions that influence both the supply of organic matter substrates and the microorganisms performing the transformations. A detailed molecular characterization of the organic matter, microbial proteomics, and the pathways and transformations by which organic matter is decomposed will be necessary to predict the direction and magnitude of the effects of environmental changes. This article describes a methodological throughput for comprehensive metabolite characterization in a single sample by direct injection Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), gas chromatography mass spectrometry (GC-MS), nuclear magnetic resonance (NMR) spectroscopy, liquid chromatography mass spectrometry (LC-MS), and proteomics analysis. This approach results in a fully-paired dataset which improves statistical confidence for inferring pathways of organic matter decomposition, the resulting CO2 and CH4 production rates, and their responses to environmental perturbation. Herein we present results of applying this method to NOM samples collected from peatlands; however, the protocol is applicable to any NOM sample (e.g., peat, forested soils, marine sediments, etc.).

This protocol describes a single throughput for complementary analytical and omics techniques culminating in a fully-paired characterization of natural organic matter and microbial proteomics in different ecosystems. This approach permits robust comparisons for identifying metabolic pathways and transformations important for describing greenhouse gas production and predicting responses to environmental change.

Singolo-velocità effettiva complementari ad alta risoluzione tecniche analitiche per la caratterizzazione di miscele complesse di sostanza organica naturale