Name: high-throughput screening for protein-based inheritance in s. cerevisiae
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Description: Watch this Scientific Journal Video about High-throughput Screening for Protein-based Inheritance in S. cerevisiae at JoVE.com

Vi tackar Sohini Chakrabortee, Sandra Jones, David Garcia, Bhupinder Bhullar, Amelia Chang, Richard She och Susan Lindquist för deras hjälp för att utveckla analyserna används i detta dokument, liksom granskarna för sina tankeväckande kommentarer.

1. Initial Overexpression
<ol>
	<li>Transform the yeast cells (in this case, BY4741 MAT a haploids) previously grown in YPD liquid (10 g of yeast extract, 20 g of peptone, and 20 g of glucose per 1 L) with the desired candidate constructs from the FLEXGene ORFeome library (yeast ORFs under the control of a galactose-inducible promoter in the URA3-marked centromericplasmid backbone, pBY01129).</li>
	<li>Use autoclaved toothpicks to pick four separate colonies from these...

<ol>
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Y., 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=Prion-inducing+domain+2-114+of+yeast+Sup35+protein+transforms+in+vitro+into+amyloid-like+filaments.">Prion-inducing domain 2-114 of yeast Sup35 protein transforms in vitro into amyloid-like filaments.</a> Proc Natl Acad Sci U S A. 94 (13), 6618-6622 (1997).</li><li>Shorter, J., Lindquist, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hsp104+catalyzes+formation+and+elimination+of+self-replicating+Sup35+prion+conformers.">Hsp104 catalyzes formation and elimination of self-replicating Sup35 prion conformers.</a> Science. 304 (5678), 1793-1797 (2004).</li><li>Ferreira, P. C., Ness, F., Edwards, S. R., Cox, B. S., Tuite, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+elimination+of+the+yeast+[PSI+]+prion+by+guanidine+hydrochloride+is+the+result+of+Hsp104+inactivation.">The elimination of the yeast [PSI+] prion by guanidine hydrochloride is the result of Hsp104 inactivation.</a> Mol Microbiol. 40 (6), 1357-1369 (2001).</li><li>Caudron, F., Barral, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+super-assembly+of+Whi3+encodes+memory+of+deceptive+encounters+by+single+cells+during+yeast+courtship.">A super-assembly of Whi3 encodes memory of deceptive encounters by single cells during yeast courtship.</a> Cell. 155 (6), 1244-1257 (2013).</li><li>Wickner, R. B., Edskes, H. K., Shewmaker, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+to+find+a+prion:+[URE3],+[PSI+]+and+[beta].">How to find a prion: [URE3], [PSI+] and [beta].</a> Methods. 39 (1), 3-8 (2006).</li><li>Chernoff, Y. O., Derkach, I. L., Inge-Vechtomov, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multicopy+SUP35+gene+induces+de-novo+appearance+of+psi-like+factors+in+the+yeast+Saccharomyces+cerevisiae.">Multicopy SUP35 gene induces de-novo appearance of psi-like factors in the yeast Saccharomyces cerevisiae.</a> Curr Genet. 24 (3), 268-270 (1993).</li><li>Brown, J. C., Lindquist, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+heritable+switch+in+carbon+source+utilization+driven+by+an+unusual+yeast+prion.">A heritable switch in carbon source utilization driven by an unusual yeast prion.</a> Genes Dev. 23 (19), 2320-2332 (2009).</li><li>Coustou, V., Deleu, C., Saupe, S., Begueret, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+protein+product+of+the+het-s+heterokaryon+incompatibility+gene+of+the+fungus+Podospora+anserina+behaves+as+a+prion+analog.">The protein product of the het-s heterokaryon incompatibility gene of the fungus Podospora anserina behaves as a prion analog.</a> Proc Natl Acad Sci U S A. 94 (18), 9773-9778 (1997).</li><li>Sopko, 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=Mapping+pathways+and+phenotypes+by+systematic+gene+overexpression.">Mapping pathways and phenotypes by systematic gene overexpression.</a> Mol Cell. 21 (3), 319-330 (2006).</li><li>Chakrabortee, 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=Intrinsically+Disordered+Proteins+Drive+Emergence+and+Inheritance+of+Biological+Traits.">Intrinsically Disordered Proteins Drive Emergence and Inheritance of Biological Traits.</a> Cell. 167 (2), 369-381 (2016).</li><li>Hu, Y., 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=Approaching+a+complete+repository+of+sequence-verified+protein-encoding+clones+for+Saccharomyces+cerevisiae.">Approaching a complete repository of sequence-verified protein-encoding clones for Saccharomyces cerevisiae.</a> Genome Res. 17 (4), 536-543 (2007).</li><li>Swain, P. 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=Inferring+time+derivatives+including+cell+growth+rates+using+Gaussian+processes.">Inferring time derivatives including cell growth rates using Gaussian processes.</a> Nat Commun. 7, 13766 (2016).</li><li>Jarosz, D. F., Lancaster, A. K., Brown, J. C., Lindquist, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+evolutionarily+conserved+prion-like+element+converts+wild+fungi+from+metabolic+specialists+to+generalists.">An evolutionarily conserved prion-like element converts wild fungi from metabolic specialists to generalists.</a> Cell. 158 (5), 1072-1082 (2014).</li><li>Neiman, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sporulation+in+the+budding+yeast+Saccharomyces+cerevisiae.">Sporulation in the budding yeast Saccharomyces cerevisiae.</a> Genetics. 189 (3), 737-765 (2011).</li><li>Gietz, D., St Jean, A., Woods, R. A., Schiestl, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+method+for+high+efficiency+transformation+of+intact+yeast+cells.">Improved method for high efficiency transformation of intact yeast cells.</a> Nucleic Acids Res. 20 (6), 1425 (1992).</li><li>Formosa, T., Nittis, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Suppressors+of+the+temperature+sensitivity+of+DNA+polymerase+alpha+mutations+in+Saccharomyces+cerevisiae.">Suppressors of the temperature sensitivity of DNA polymerase alpha mutations in Saccharomyces cerevisiae.</a> Mol Gen Genet. 257 (4), 461-468 (1998).</li><li>Christiano, R., Nagaraj, N., Frohlich, F., Walther, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+proteome+turnover+analyses+of+the+Yeasts+S.+cerevisiae+and+S.+pombe.">Global proteome turnover analyses of the Yeasts S. cerevisiae and S. pombe.</a> Cell Rep. 9 (5), 1959-1965 (2014).</li><li>Lancaster, A. K., Nutter-Upham, A., Lindquist, S., King, O. 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De första jäst prionerna identifierades genom sin ovanliga fenotyper och förbryllande mönster av arv. Det som kännetecknen dessa prioner användes sedan för att bygga algoritmer och beräkningsverktyg till skärmen för ytterligare prion proteiner. Den metod som beskrivs här, däremot är experimentell och bygger på övergående överuttryck att skapa en varaktig förändring-en stabil stat-kodade i protein konformation. Dock om effektivitet ”sådd” prion montering av överuttryck för något visst protein ä...

Biologiska system upplever ofta övergående fluktuationer i protein överflöd. Det är fortfarande oklart huruvida dessa har en bestående inverkan forma fenotypen av en organism eller framtida generationer. De mest kända förekomsterna av detta biologi innebär en sällsynt klass av proteiner, prioner, som driver framväxten av ärftliga egenskaper utan genomet modifiering. I stället överföra dessa proteinaceous och isjukdomar partiklar fenotyper via självförstärkande ändringar till protein konformation1,2. Denna typ av arv upptäcktes som orsaken till de ovanliga nedärvningsmönster av en förödande neurodegenerativ sjukdom. Studier i organismer som sträcker sig från svampar till däggdjur3,4,5,6,7,8,9,10 har sedan visade emellertid att prion-liknande element kan ge adaptiva värde. Prioner har dock setts som en fascinerande men sällsynt biologiska oddity.
Detta rådande visdom hålls delvis eftersom karakterisering av proteinbaserade arv har länge begränsats av ett litet antal exempel. Senaste systematisk screening ansträngningar har breddat bilden betydligt genom att identifiera flera nya bona fide prioner11 och nästan två dussin protein domäner12 med kapacitet att bränsle prion-liknande konfirmerande konvertering. Men eftersom dessa metoder har allmänhet fokuserade på stark amino syra ordnar fördomar, dela prionerna som har identifierats biokemiska egenskaperna för den grundande jäst prioner [PSI+]13,14, [URE3]15och [RNQ+]11,16. Dessa inkluderar: (1) modulära domäner som är rika på långa polymera sträckor av asparagin (N) och glutamin (Q), (2) församlingen in i en amyloid [PRION+] konformation17,18,19, och 3) fullständig tillit till disaggregase Hsp104 funktion för trogna förökning från mor till dotter13,20,21. Faktiskt, många bona fide prioner, inklusive [GAR+], [Het-s], och även den ursprungliga prionen (PrPSc), skulle missas under sådana stränga kriterier. Kanske ännu viktigare, skulle de kunna fånga några nya mekanismer av proteinbaserade arv22. Sanna biologiska vidden av sådana fenomen kan således vara betydligt vanligare i naturen än man tidigare antagit.
För att undersöka denna fråga, var en hög genomströmning, proteomet-omfattande strategi anställd. Ett kännetecken för alla prioner, inklusive PrPSc, [GAR+], och [Het-s], är att de övergående överuttryck av kausala proteinerna starkt ökar graden av prion förvärv15,23,24,25,26. Vi drog fördel av denna funktion att systematiskt fråga, i hela jäst ORFeome, om stabil protein-baserade, epigenetiska stater kan inledas av övergående förmå överuttryck av enskilda proteiner. Det är välkänt att protein överuttryck kan förändra fenotyper27. Prion proteiner är dock ovanligt eftersom deras tillfällig överproduktion ger en förändring i fenotyp som är ärftligt för många hundratals generationer efter den inledande överuttryck. Tidigare tog vi fördel av denna funktion, liksom de ovanliga nedärvningsmönster proteinbaserade genetiska element, för att identifiera dussintals proteiner som klarar av heritably nytt kablage fenotypiska landskap utan att ändra i genomet28. Även om några identifierade var proteiner tidigare känt som prioner, de flesta var inte, betonar kraften i denna strategi att avslöja nya former av protein-baserade arv.

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			<td height="21" style="width:356px;height:21px;">Guanidine hydrochloride</td>
			<td style="width:299px;">Sigma</td>
			<td style="width:125px;">Cat#G3272-25G</td>
			<td style="width:179px;">Chemical</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Manganese chloride</td>
			<td>Sigma</td>
			<td>Cat#M8054-100G</td>
			<td>Chemical</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Ethidium bromide</td>
			<td>Sigma</td>
			<td>E1510</td>
			<td>Chemical</td>
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		<tr height="21">
			<td height="21" style="height:21px;">5-Fluoroorotic Acid</td>
			<td>Sigma</td>
			<td>Cat#F5013-50MG</td>
			<td>Chemical</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">BY4741 MATa (his3&#916;1 leu2&#916;0 LYS2 met15&#916;0 ura3&#916;0)</td>
			<td>Winston et al., 1995; Brachmann et al., 1998</td>
			<td>N/A</td>
			<td>Yeast strain</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">BY4741 MAT&#945; (his3&#916;1 leu2&#916;0 lys2&#916;0 MET15 ura3&#916;0)</td>
			<td>Winston et al., 1995; Brachmann et al., 1998</td>
			<td>N/A</td>
			<td>Yeast strain</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Hsp70 (K69M)&#160;</td>
			<td>Jarosz et al., 2014b</td>
			<td>N/A</td>
			<td>Plasmid</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FLEXGene library</td>
			<td>Hu et al., 2007</td>
			<td>N/A</td>
			<td>Plasmid library</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dextrose (glucose)</td>
			<td>Fisher Scientific</td>
			<td>D16-3</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Raffinose</td>
			<td>Sigma</td>
			<td>R0250-25G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Galactose</td>
			<td>Fisher Scientific</td>
			<td>BP656-500</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CSM</td>
			<td>Sunrise Science</td>
			<td>1001-100</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CSM-URA</td>
			<td>Sunrise Science</td>
			<td>1004-100</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CSM-LYS</td>
			<td>Sunrise Science</td>
			<td>1032-100</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CSM-MET</td>
			<td>Sunrise Science</td>
			<td>1019-100</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">CSM-LYS-MET</td>
			<td>Sunrise Science</td>
			<td>1035-100</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">yeast extract</td>
			<td>Fisher Scientific</td>
			<td>BP1422-2</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">peptone</td>
			<td>Research Products International</td>
			<td>P20240-5000</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">bacto-peptone</td>
			<td>BD</td>
			<td align="right">211677</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">glycerol</td>
			<td>EMD Millipore</td>
			<td>GX0185-2</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">yeast nitrogen base w/o amino acids</td>
			<td>BD</td>
			<td align="right">291920</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">agar</td>
			<td>IBI Scientific</td>
			<td>IB49172</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Adenine sulfate</td>
			<td>Sigma</td>
			<td>A3159-25G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Potassium acetate</td>
			<td>Sigma</td>
			<td>P1190-500G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Uracil</td>
			<td>Sigma</td>
			<td>U0750-100G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Histidine</td>
			<td>Sigma</td>
			<td>H8000-100G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Leucine</td>
			<td>Sigma</td>
			<td>L8000-25G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Lysine</td>
			<td>Sigma</td>
			<td>L5501-25G</td>
			<td>Media component</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">RNase I&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>EN0601</td>
			<td>Enzyme</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">biotinylated DNase</td>
			<td>Thermo Fisher Scientific</td>
			<td>AM1906</td>
			<td>Enzyme</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">zymolyase 100T (yeast lytic enzyme)</td>
			<td>Sunrise Science</td>
			<td>N0766555</td>
			<td>Enzyme</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Microplate reader</td>
			<td>BioTek</td>
			<td>Synergy H1</td>
			<td>Equipment</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Microplate stacker</td>
			<td>BioTek</td>
			<td>BioStack3</td>
			<td>Equipment</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Plate filler</td>
			<td>BiotTek</td>
			<td>EL406</td>
			<td>Equipment</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Liquid handling robot</td>
			<td>Beckman Coulter</td>
			<td>Biomek FX</td>
			<td>Equipment</td>
		</tr>
	</tbody>
</table>

high-throughput screening for protein-based inheritance in s. cerevisiae

Kodning av biologisk information som är tillgänglig för framtida generationer uppnås generellt via förändringar i DNA-sekvensen. Långlivat arv kodade i protein konformation (snarare än sekvens) har länge setts som paradigm-shifting men sällsynt. De bästa kännetecknas exemplen på sådana epigenetiska faktorer är prioner, som äger en egen montering beteende som kan driva den ärftliga manifestationen av nya fenotyper. Många arketypiska prioner Visa en slående N/Q-rika sekvens bias och montera in en amyloid fållan. Dessa ovanliga funktioner har informerat de flesta screening ansträngningar att identifiera nya prion proteiner. Minst tre kända prioner (inklusive grundande prioner, PrPSc) dock inte hysa dessa biokemiska egenskaper. Vi har därför utvecklat en alternativ metod för att undersöka omfattningen av proteinbaserade arv baserat på en egenskap av massa åtgärder: den övergående överuttryck av prion proteiner ökar frekvensen vid vilken de förvärvar en self-mallhantering konformation. Detta dokument beskriver en metod för att analysera jästen ORFeome förmåga att framkalla proteinbaserade arv. Använda denna strategi, fann vi tidigare att &gt; 1% jäst proteiner kunde bränsle uppkomsten av biologiska egenskaper som var långlivat, stabil, och uppstod oftare än genetisk mutation. Detta tillvägagångssätt kan vara anställd i hög genomströmning över hela ORFeomes eller som en riktad screening paradigm för specifika genetiska nätverk eller stimuli från omgivningen. Precis som framåt genetiska skärmar definiera många utvecklings- och signalering vägar, ger dessa tekniker en metod för att undersöka påverkan av proteinbaserade arv i biologiska processer.

Protein overexpression is known to dramatically alter cellular phenotypes27. Indeed, with an initial screening approach, hundreds of new phenotypes were reproducibly recovered from the overexpression of clones from the yeast ORFeome using just ten stressors. However, the assays described above allow for the assessment of whether cells retain any long-term stable phenotypes following this overexpression. One protein capable of encoding such a state is Psp1. Psp1 is ...

Watch this Scientific Journal Video about High-throughput Screening for Protein-based Inheritance in S. cerevisiae at JoVE.com

High-throughput Screening for Protein-based Inheritance in S. cerevisiae

Det här protokollet beskriver en hög genomströmning metod att funktionellt skärmen för protein-baserade arv i S. cerevisiae.

High-throughput Screening för Protein-baserade arv i S. cerevisiae

The encoding of biological information that is accessible to future generations is generally achieved via changes to the DNA sequence. Long-lived ...

Research

JoVE Journal

Biology

High-throughput Screening for Protein-based Inheritance in S. cerevisiae

Assay for Adhesion and Agar Invasion in S. cerevisiae

Analys för vidhäftningsförmåga och agar Invasion i S. cerevisiae

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, ...

A high-throughput method to globally study the organelle morphology in S. cerevisiae

En hög genomströmning metod för att globalt studera organell morfologi i S. cerevisiae

High-throughput methods to examine protein localization or organelle morphology is an effective tool for studying protein interactions and can help ...

Identification of protein complexes with quantitative proteomics in S. cerevisiae

Kartläggning av protein komplex med kvantitativa proteomik i S. cerevisiae

Lipids are the building blocks of cellular membranes that function as barriers and in compartmentalization of cellular processes, and recently, as ...

High-throughput Screening and Biosensing with Fluorescent C. elegans Strains

High-throughput Screening and Biosensing with Fluorescent C. elegans Strains

High-throughput screening och biosensing med fluorescerande C. elegans Stammar

High-throughput screening (HTS) is a powerful approach for identifying chemical modulators of biological processes.  However, many compounds identified in ...

High Throughput Screening of Fungal Endoglucanase Activity in Escherichia coli

High Throughput Screening of Fungal Endoglucanase Activity in Escherichia coli

Hög throughput screening av svamp Endoglucanase aktivitet i Escherichia coli</em

Cellulase enzymes (endoglucanases, cellobiohydrolases, and &beta;-glucosidases) hydrolyze cellulose into component sugars, which in turn can be converted ...

Flow Cytometry-based Purification of S. cerevisiae Zygotes

Flow Cytometry-based Purification of S. cerevisiae Zygotes

Flödescytometri-baserad rening av<em&gt; S. cerevisiae</em&gt; Zygoter

Zygotes are essential intermediates between haploid and diploid states in the life cycle of many organisms, including yeast (Figure 1) 1. S. cerevisiae ...

High-throughput Screening for Small-molecule Modulators of Inward Rectifier Potassium Channels

High-throughput screening för små molekyler Modulatorer för aktiv kanaler Likriktare Kalium

Specific members of the inward rectifier potassium (Kir) channel family are postulated drug targets for a variety of disorders, including hypertension, ...

High-throughput Functional Screening using a Homemade Dual-glow Luciferase Assay

Hög kapacitet Funktionell Screening med hjälp av en hemmagjord Dual-glöd luciferas analys

We present a rapid and inexpensive high-throughput screening protocol to identify transcriptional regulators of alpha-synuclein, a gene associated with ...

High Throughput Quantitative Expression Screening and Purification Applied to Recombinant Disulfide-rich Venom Proteins Produced in E. coli

High Throughput Quantitative Expression Screening and Purification Applied to Recombinant Disulfide-rich Venom Proteins Produced in E. coli

Hög genomströmning Kvantitativ Expression Screening och rening Tillämpat på rekombinant Disulfide rika Venom proteiner som produceras i<em&gt; E. coli</em

Escherichia coli (E. coli) is the most widely used expression system for the production of recombinant proteins for structural and functional studies. ...

High-throughput Screening for Chemical Modulators of Post-transcriptionally Regulated Genes

Hög kapacitet Screening för kemiska modulatorer av Post-transkription Reglerade Gener

Both transcriptional and post-transcriptional regulation have a profound impact on genes expression. However, commonly adopted cell-based screening assays ...