Name: investigating the detrimental effects of low pressure plasma sterilization on the survival of bacillus subtilis spores using live cell microscopy
Uploaded: 2017-11-30T14:00:00
Description: Watch this Scientific Journal Video about Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy at JoVE.com

Forfatterne tak Andrea Schröder for hendes fremragende teknisk bistand under dele af dette arbejde og Nikea J. Ulrich for hendes hjælp under video-optagelser. Vi vil også gerne takke Lyle A. Simmons for sin generøse donation af Bacillus subtilis stammer: LAS72 og LAS24. Dette arbejde blev støttet i dele af tilskud fra den tyske Research Foundation (DFG) Paketantrag (PlasmaDecon PAK 728) til PA (AW 7/3-1) og RM (MO 2023/2-1) og DLR yder DLR-FuW-Projekt ISS liv, Programm RF-FuW, Teilprogramm 475 (til F.M.F, M.R. og R.M.). F.M.F. blev støttet af et ph.d.-stipendium af Helmholtz plads Life Sciences Research School (SpaceLife) på tysk Aerospace Center (DLR) i Köln, Tyskland, som blev finansieret af Helmholtz Association (Helmholtz-Gemeinschaft) over en periode på seks år ( Grant nr. VH-ko-300) og modtog yderligere midler fra DLR, herunder Aerospace direktionen og flyvemedicin Institut. Resultaterne af denne undersøgelse vil indgå i Ph.D.-afhandling af Felix M. Fuchs.

1. bacillus subtilis Spore produktion og rensning
<ol><li>Overfør en 5 mL overnight kultur af de respektive for at spore produktionen, B. subtilis stamme, suppleret med passende antibiotika til 200 mL dobbelt-styrke flydende Schaeffer sporulering medium (pr. liter 16 g næringsstof bouillon, KCl 2 g, 0,5 g MgSO 4* 7 H2O, 2 mL 1 M Ca (3)2, 2 mL 0,1 M frihedsbevægelse2 * • 4 H2O, 2 mL 1 mM FeSO4, 2 mL 50% (w/v) glukose<sup ...

<ol>
	<li>Nicholson, W. L., Schuerger, A. C., Race, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Migrating+microbes+and+planetary+protection.">Migrating microbes and planetary protection.</a> Trends Microbiol. 17, 389-392 (2009).</li><li>COSPAR. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=COSPAR+Planetery+Protection+Policy.">COSPAR Planetery Protection Policy.</a> Space Research Today, COSPAR's Information Bulletin. 193, 1-14 (2015).</li><li>De Geyter, N., Morent, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonthermal+plasma+sterilization+of+living+and+nonliving+surfaces.">Nonthermal plasma sterilization of living and nonliving surfaces.</a> Annu Rev Biomed Eng. 14, 255-274 (2012).</li><li>Shimizu, S., 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=Cold+atmospheric+plasma+-+A+new+technology+for+spacecraft+component+decontamination.">Cold atmospheric plasma - A new technology for spacecraft component decontamination.</a> Planet. Space Sci. 90, 60-71 (2014).</li><li>Lerouge, S., Fozza, A. C., Wertheimer, M. R., Marchand, R., Yahia, L. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sterilization+by+Low-Pressure+Plasma:+The+Role+of+Vacuum-Ultraviolet+Radiation.">Sterilization by Low-Pressure Plasma: The Role of Vacuum-Ultraviolet Radiation.</a> Plasma Polym. 5, 31-46 (2000).</li><li>Rossi, F., Kyli&#225;n, O., Rauscher, H., Gilliland, D., Sirghi, L. <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+low-pressure+plasma+discharge+for+the+decontamination+and+sterilization+of+medical+devices.">Use of a low-pressure plasma discharge for the decontamination and sterilization of medical devices.</a> Pure Appl. Chem. 80, 1939-1951 (2008).</li><li>Halfmann, H., Hauser, J., Awakowicz, P., Koller, M., Esenwein, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+double+inductively+coupled+low-pressure+plasma+for+sterilization+of+medical+implant+materials.">A double inductively coupled low-pressure plasma for sterilization of medical implant materials.</a> Biomed Tech (Berl). 53, 199-203 (2008).</li><li>Halfmann, H., Denis, B., Bibinov, N., Wunderlich, J., Awakowicz, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+the+most+efficient+VUV/UV+radiation+for+plasma+based+inactivation+of+Bacillus+atrophaeus+spores.">Identification of the most efficient VUV/UV radiation for plasma based inactivation of Bacillus atrophaeus spores.</a> J. Phys. D: Appl. Phys. 40, 5907 (2007).</li><li>Vaishampayan, P., 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=Bacillus+horneckiae+sp.+nov.,+isolated+from+a+spacecraft-assembly+clean+room.">Bacillus horneckiae sp. nov., isolated from a spacecraft-assembly clean room.</a> Int J Syst Evol Microbiol. 60, 1031-1037 (2010).</li><li>Mandic-Mulec, I., Stefanic, P., van Elsas, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ecology+of+Bacillaceae.">Ecology of Bacillaceae.</a> Microbiol Spectr. 3, (2015).</li><li>Alekhova, T. A., 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=Diversity+of+bacteria+of+the+genus+Bacillus+on+board+of+international+space+station.">Diversity of bacteria of the genus Bacillus on board of international space station.</a> Dokl Biochem Biophys. 465, 347-350 (2015).</li><li>Claus, D., Bekerley, R. C. W., Sneath, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genus+Bacillus+Cohn+1872.">Genus Bacillus Cohn 1872.</a> Bergey's manual of systematic bacteriology. 2, 1105-1141 (1986).</li><li>Setlow, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spore+Resistance+Properties.">Spore Resistance Properties.</a> Microbiol Spectr. 2, (2014).</li><li>Setlow, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spores+of+Bacillus+subtilis:+their+resistance+to+and+killing+by+radiation,+heat+and+chemicals.">Spores of Bacillus subtilis: their resistance to and killing by radiation, heat and chemicals.</a> J Appl Microbiol. 101, 514-525 (2006).</li><li>Roth, S., Feichtinger, J., Hertel, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+Bacillus+subtilis+spore+inactivation+in+low-pressure,+low-temperature+gas+plasma+sterilization+processes.">Characterization of Bacillus subtilis spore inactivation in low-pressure, low-temperature gas plasma sterilization processes.</a> J Appl Microbiol. 108, 521-531 (2010).</li><li>Roth, S., Feichtinger, J., Hertel, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Response+of+Deinococcus+radiodurans+to+low-pressure+low-temperature+plasma+sterilization+processes.">Response of Deinococcus radiodurans to low-pressure low-temperature plasma sterilization processes.</a> J Appl Microbiol. 109, 1521-1530 (2010).</li><li>Setlow, B., Setlow, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+DNA+repair+in+Bacillus+subtilis+spore+resistance.">Role of DNA repair in Bacillus subtilis spore resistance.</a> J Bacteriol. 178, 3486-3495 (1996).</li><li>Schaeffer, P., Millet, J., Aubert, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Catabolic+repression+of+bacterial+sporulation.">Catabolic repression of bacterial sporulation.</a> Proc. Natl. Acad. Sci. 54, 704-711 (1965).</li><li>Simmons, L. A., 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=Comparison+of+responses+to+double-strand+breaks+between+Escherichia+coli+and+Bacillus+subtilis+reveals+different+requirements+for+SOS+induction.">Comparison of responses to double-strand breaks between Escherichia coli and Bacillus subtilis reveals different requirements for SOS induction.</a> J Bacteriol. 191, 1152-1161 (2009).</li><li>Raguse, M., 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=Improvement+of+Biological+Indicators+by+Uniformly+Distributing+Bacillus+subtilis+Spores+in+Monolayers+To+Evaluate+Enhanced+Spore+Decontamination+Technologies.">Improvement of Biological Indicators by Uniformly Distributing Bacillus subtilis Spores in Monolayers To Evaluate Enhanced Spore Decontamination Technologies.</a> Appl Environ Microbiol. 82, 2031-2038 (2016).</li><li>Horneck, G., 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=Protection+of+bacterial+spores+in+space,+a+contribution+to+the+discussion+on+Panspermia.">Protection of bacterial spores in space, a contribution to the discussion on Panspermia.</a> Orig Life Evol Biosph. 31, 527-547 (2001).</li><li>Opretzka, J., Benedikt, J., Awakowicz, P., Wunderlich, J., Keudell, A. v. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+chemical+sputtering+during+plasma+sterilization+of+Bacillus+atrophaeus.">The role of chemical sputtering during plasma sterilization of Bacillus atrophaeus.</a> J. Phys. D: Appl. Phys. 40, 2826 (2007).</li><li>Stapelmann, K., 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=Utilization+of+low-pressure+plasma+to+inactivate+bacterial+spores+on+stainless+steel+screws.">Utilization of low-pressure plasma to inactivate bacterial spores on stainless steel screws.</a> Int. J. Astrobiol. 13, 597-606 (2013).</li><li>Raguse, M., 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=Understanding+of+the+importance+of+the+spore+coat+structure+and+pigmentation+in+the+Bacillus+subtilis+spore+resistance+to+low-pressure+plasma+sterilization.">Understanding of the importance of the spore coat structure and pigmentation in the Bacillus subtilis spore resistance to low-pressure plasma sterilization.</a> J. Phys. D: Appl. Phys. 49, 285401 (2016).</li><li>Pandey, R., 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=Live+cell+imaging+of+germination+and+outgrowth+of+individual+Bacillus+subtilis+spores;+the+effect+of+heat+stress+quantitatively+analyzed+with+SporeTracker.">Live cell imaging of germination and outgrowth of individual Bacillus subtilis spores; the effect of heat stress quantitatively analyzed with SporeTracker.</a> PloS one. 8, e58972 (2013).</li></ol>

Sterilisation af overflader ved hjælp af lav-temperatur, lavtryks plasma er en lovende alternativ til temmelig konventionelle sterilisation procedurer såsom behandling med ioniserende stråling, kemikalier (f.eks. gasser som H2O2 eller ethylenoxid) eller tørre og fugtige varme23. Almindelig sterilisation metoder for det meste give en effektiv sterilisering, men de er kendt for at påvirke det behandlede materiale og udgør en potentiel risiko for operatoren. Lavtr...

Et vigtigt mål for udforskning af rummet er søgning efter underskrifter af livsformer og biomolekyler på andre planeter og måner i vores solsystem. Overførsel af mikroorganismer eller biomolekyler af jordbaserede oprindelse til kritiske områder for efterforskning er i særlig risiko for indvirkning på udvikling og integritet af liv-påvisning missioner på planeter såsom Mars og Europa1. De internationale retningslinjer for planeternes beskyttelse, etableret af Udvalget for Space forskning (COSPAR) i 1967, pålægge strenge regler på robotic og bemandede missioner til andre planeter, deres måner, asteroider og andre himmellegemer og regulere den rengøring og sterilisering af et rumfartøj og kritiske hardwarekomponenter forudgående at iværksætte for at eliminere forurener jordbaserede mikroorganismer og forhindre krydskontaminering af himmellegemer2. I det sidste årti, har anvendelsen af ikke-termiske plasmaer fået bred opmærksomhed i biomedicinske og ernæringsmæssige forskning samt i rumflyvning programmer3,4,5. Plasma Sterilisation er en lovende alternativ til konventionelle sterilisation metoder, som det tilbyder hurtig og effektiv mikrobielle inaktivering6, samtidig være blid til følsomme og varme labile materialer. Plasma udledninger indeholder en blanding af reaktive agenter som frie radikaler, ladede partikler, neutral/ophidset atomer, fotoner i ultraviolet (UV) og vakuum ultraviolet (VUV) spektrum, som fører til hurtige mikrobielle inaktivering3. I denne undersøgelse bruger vi lavtryks plasma genereret af dobbelt Induktivt koblet lavtryks plasma (DICP) kilde7,8 til at inaktivere Bacillus subtilis endosporer fordelt på glasoverfladen test.
Gram-positive bakterier af familien Bacillaceae er vidt udbredt i naturlige habitater fra jord, sedimenter og luften, usædvanlige miljøer såsom renrums faciliteter og den internationale rumstation9,10 ,11. De mest forskellige træk ved slægten Bacillus er evnen til at danne meget resistente hvilende endosporer (herefter benævnt sporer) for at overleve ugunstige betingelser, såsom næringsstoffer udtynding12. Sporerne er generelt mere resistente end deres vegetative celle kolleger til en bred vifte af behandlinger og miljøbelastninger, herunder varme, UV-, gammastråling, udtørring, mekanisk afbrydelse og giftige kemikalier, såsom stærke iltningsmidler eller ændring af pH agenter (gennemgik i referencer13,14) og er derfor ideelle objekter for at teste effektiviteten af mikrobielle inaktivering metoder. Da genomisk DNA er et større mål af plasma behandling af bakterier15,16, reparation af plasma-induceret DNA læsioner (f.eks. DNA dobbeltstrenget pauser) efter spore genoplivning er afgørende for overlevelse af bakterier13, 17.
Dermed, vi studerer spireevne af sporer og rollen af DNA reparation under sporespiring og udvækst efter behandling af sporer med lavt tryk argon plasma af følgende individuelle sporer og deres udtryk for fluorescens-mærket DNA reparation protein RecA med tidsopløst Konfokal Fluorescens mikroskopi. Vi giver en trinvis instruktion i forberedelsen af B. subtilis sporer i encellelag for at opnå reproducerbare testresultater, behandling af spore encellelag med lavt tryk plasma for sterilisation, forberedelse af plasma behandlet sporer for ultrastrukturelle evaluering ved hjælp af scanning elektronmikroskopi (SEM), og levende celle mikroskopi analyse i forbindelse med individuelle sporer i koncert med overvågning af den aktive DNA reparation processer, der forekommer inden for cellen i svar til plasma behandling.

<table>
	<tbody>
		<tr>
			<td>Two substance nozzle (model 970-8)</td>
			<td>Schlick</td>
			<td>14,404</td>
			<td>230 V, 50 Hz, D 4.484/8, 0.8 mm bore diameter</td>
		</tr>
		<tr>
			<td>Luria Bertani Medium</td>
			<td>Sigma Aldrich</td>
			<td>70122-100G</td>
			<td></td>
		</tr>
		<tr>
			<td>Tube connectors</td>
			<td>Festo</td>
			<td>n/a</td>
			<td>G 1/8</td>
		</tr>
		<tr>
			<td>Magnetvalve DO35-3/2NC-G018-230AC</td>
			<td>Bosch Rexroth</td>
			<td>820005100</td>
			<td></td>
		</tr>
		<tr>
			<td>PLN Polyamid tube</td>
			<td>Festo</td>
			<td>558206</td>
			<td>d = 6 mm</td>
		</tr>
		<tr>
			<td>Glass slides</td>
			<td>VWR</td>
			<td>48300-026</td>
			<td></td>
		</tr>
		<tr>
			<td>Electric Timer 550-2-C</td>
			<td>Gefran</td>
			<td>F000074</td>
			<td>220 V</td>
		</tr>
		<tr>
			<td>attofluor cell chamber</td>
			<td>Menzel, Fisher Ref.</td>
			<td>3406816</td>
			<td>d=25 mm, round</td>
		</tr>
		<tr>
			<td>MgSO4*7 H2O</td>
			<td>Sigma Aldrich</td>
			<td>13152</td>
			<td></td>
		</tr>
		<tr>
			<td>Ca(NO3)2</td>
			<td>Sigma Aldrich</td>
			<td>202967</td>
			<td></td>
		</tr>
		<tr>
			<td>MnCl2 * 4 H2O</td>
			<td>Sigma Aldrich</td>
			<td>244589</td>
			<td></td>
		</tr>
		<tr>
			<td>FeSO4 * 7H2O</td>
			<td>AppliChem</td>
			<td>13446-34-9</td>
			<td></td>
		</tr>
		<tr>
			<td>Glucose</td>
			<td>Merck</td>
			<td>215422</td>
			<td></td>
		</tr>
		<tr>
			<td>KCl</td>
			<td>Sigma Aldrich</td>
			<td>P9541-500G</td>
			<td></td>
		</tr>
		<tr>
			<td>Nutrient Broth (NB)</td>
			<td>Merck</td>
			<td>105443</td>
			<td></td>
		</tr>
		<tr>
			<td>Luria-Bertani (LB)</td>
			<td>Merck</td>
			<td>110283</td>
			<td></td>
		</tr>
		<tr>
			<td>96-wellplate</td>
			<td>ThermoFisher</td>
			<td>243656</td>
			<td></td>
		</tr>
		<tr>
			<td>Zeiss LSM 780, Axio Observer Z1</td>
			<td>Carl Zeiss Microscopy GmbH</td>
			<td>n/a</td>
			<td></td>
		</tr>
		<tr>
			<td>Leo 1530 Gemini</td>
			<td>Carl Zeiss Microscopy GmbH</td>
			<td>n/a</td>
			<td></td>
		</tr>
		<tr>
			<td>ZEN 2 and ZEN lite 2012 (Software)</td>
			<td>Carl Zeiss Microscopy GmbH</td>
			<td>n/a</td>
			<td></td>
		</tr>
		<tr>
			<td>SigmaPlot, version 13.0 (Statistic software)</td>
			<td>Systat GmbH, Erkrath, Germany</td>
			<td>n/a</td>
			<td></td>
		</tr>
		<tr>
			<td>Attofluor cell chamber</td>
			<td>Invitrogen</td>
			<td>A7816</td>
			<td></td>
		</tr>
		<tr>
			<td>&#181;-Dish 35 mm, high Grid-500 Glass Bottom</td>
			<td>ibidi</td>
			<td>81168</td>
			<td></td>
		</tr>
	</tbody>
</table>

investigating the detrimental effects of low pressure plasma sterilization on the survival of bacillus subtilis spores using live cell microscopy

Plasma Sterilisation er en lovende alternativ til konventionelle sterilisation metoder til industriel, klinisk, og rumflyvninger formål. Lavtryk plasma (LPP) udledninger indeholder et bredt spektrum af aktive arter, som fører til hurtige mikrobielle inaktivering. For at undersøge effektiviteten og mekanismer for Sterilisation af LPP, bruger vi sporer af testorganismen Bacillus subtilis på grund af deres ekstraordinære modstand mod konventionelle sterilisation procedurer. Vi beskrive produktionen af B. subtilis spore encellelag, sterilisering af lavtryk plasma i en dobbelt Induktivt koblet plasma-reaktoren, karakterisering af spore morfologi ved hjælp af scanning elektronmikroskopi (SEM), og den analyse af spiring og udvækst af sporer af levende celle mikroskopi. Et større mål af plasma arter er genomisk materiale (DNA) og reparation af plasma-induceret DNA læsioner på spore genoplivning er afgørende for overlevelse af organismen. Her, vi studerer spireevne af sporer og rollen af DNA reparation under sporespiring og udvækst efter behandling med LPP ved at spore fluorescently mærket DNA reparation proteiner (RecA) med tidsopløst Konfokal Fluorescens mikroskopi. Behandlet og ubehandlet spore encellelag er aktiveret for spiring og visualiseret med en inverteret Konfokal levende celle mikroskop over tid til at følge reaktionen af individuelle sporer. Vores observationer viser, at brøkdel af spire og outgrowing sporerne er afhængig af varigheden af LPP-behandling at nå minimum efter 120 s. RecA-YFP (gul fluorescens protein) Fluorescens blev opdaget kun i par sporer og udviklet i alle outgrowing celler med en lille forhøjning i LPP-behandlede sporer. Desuden, nogle af de vegetative bakterier stammer fra LPP-behandlede sporer viste en stigning i cytoplasma og en tendens til at lyse. De beskrevne metoder til analyse af enkelte sporer kunne være eksemplarisk for studiet af andre aspekter af sporespiring og udvækst.

Overlevelse af plasma-behandlede B. subtilis sporer
Plasma behandling af B. subtilis sporer anvendes i denne undersøgelse viser et fald i overlevelse med stigende varighed af plasma behandling (figur 2). Sporer af stammen at udtrykke recA-gen sammenvokset til YFP viste overlevelse kurver svarende til sporer af vildtype stamme, som angiver, at den genetiske modifikation har i...

Watch this Scientific Journal Video about Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy at JoVE.com

Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy

Undersøge de skadelige effekter af lav pres Plasma sterilisation på overlevelse af Bacillus subtilis sporer ved hjælp af Live celle mikroskopi

Denne protokol illustrerer de vigtige træk skridt forpligtet til at vurdere relevansen af overvågning vitalitet parameter og DNA reparationsprocesser i genoplive Bacillus subtilis sporer efter behandling med lavt tryk plasma med tracking Fluorescens-mærket DNA reparere proteiner via tidsopløst Fluorescens mikroskopi og scanning Elektron Mikroskopi.

Plasma sterilization is a promising alternative to conventional sterilization methods for industrial, clinical, and spaceflight purposes. Low pressure ...

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Biology

Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy

Imaging Mismatch Repair and Cellular Responses to DNA Damage in Bacillus subtilis

Imaging Mismatch Repair and Cellular Responses to DNA Damage in Bacillus subtilis

Both prokaryotes and eukaryotes respond to DNA damage through a complex set of physiological changes. Alterations in gene expression, the redistribution ...

Identifying the Effects of BRCA1 Mutations on Homologous Recombination using Cells that Express Endogenous Wild-type BRCA1

The functional analysis of missense mutations can be complicated by the presence in the cell of the endogenous protein. Structure-function analyses of the ...

FRET Microscopy for Real-time Monitoring of Signaling Events in Live Cells Using Unimolecular Biosensors

FRET mikroskopi af Real-time overvågning af signalering begivenheder i levende celler Brug unimolekylær Biosensorer

F&ouml;rster resonance energy transfer (FRET) microscopy  continues to gain increasing interest as a technique for real-time monitoring  of biochemical ...

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

Levende Imaging af Apoptotisk Cell Clearance under<em&gt; Drosophila</em&gt; Embryogenese

The  proper elimination of unwanted or aberrant cells through apoptosis and  subsequent phagocytosis (apoptotic cell clearance) is crucial for normal  ...

Analyzing Craniofacial Morphogenesis in Zebrafish Using 4D Confocal Microscopy

Analyserer kraniofacial Morfogenese i zebrafisk Brug 4D konfokal mikroskopi

Time-lapse imaging is a technique that allows for the direct observation of the process of morphogenesis, or the generation of shape. Due to their optical ...

Light Sheet Fluorescence Microscopy of Plant Roots Growing on the Surface of a Gel

Light Sheet Fluorescens mikroskopi af planterødder Dyrkning på overfladen af ​​en gel

One of the key questions in understanding plant development is how single cells behave in a larger context of the tissue. Therefore, it requires the ...

Standardized Method for High-throughput Sterilization of Arabidopsis Seeds

Standardiseret metode til høj overførselshastighed Sterilisation af Arabidopsis frø

Arabidopsis thaliana (Arabidopsis) seedlings often need to be grown on sterile media. This requires prior seed sterilization to prevent the growth of ...

Single-cell Microfluidic Analysis of Bacillus subtilis

Single-cell Microfluidic Analysis of Bacillus subtilis

Encellede mikrofluid analyse af Bacillus subtilis

Microfluidic technology overcomes many of the limitations to traditional analytical methods in microbiology. Unlike bulk-culture methods, it offers ...

Biological Sample Preparation by High-pressure Freezing, Microwave-assisted Contrast Enhancement, and Minimal Resin Embedding for Volume Imaging

Biologiske prøveforberedelse højtryks frysebehandling, mikrobølgeovn-assisteret kontrastforbedring og Minimal harpiks indlejring for volumen Imaging

The described sample preparation technique is designed to combine the best quality of ultrastructural preservation with the most suitable contrast for the ...

Live-Cell Imaging of the Life Cycle of Bacterial Predator Bdellovibrio bacteriovorus using Time-Lapse Fluorescence Microscopy

Live-Cell Imaging of the Life Cycle of Bacterial Predator Bdellovibrio bacteriovorus using Time-Lapse Fluorescence Microscopy

Live-Cell Imaging af livscyklus bakteriel Predator Bdellovibrio bacteriovorus ved hjælp af Time-Lapse Fluorescens mikroskopi

Bdellovibrio bacteriovorus is a small gram-negative, obligate predatory bacterium that kills other gram-negative bacteria, including harmful pathogens. ...