Name: horizontal gel electrophoresis for enhanced detection of protein-rna complexes
Uploaded: 2017-07-28T14:00:00
Description: Watch this Scientific Journal Video about Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes at JoVE.com

Vi takker Laura Vanderploeg for at forberede tal. Arbejde i Sheets lab understøttes af NSF grant 1050395 og NIH grant (R21HD076828). Arbejde i Ryder-labet understøttes af NIH-tilskud R01GM117237 og R01GM117008. Megan Dowdle støttes af et SciMed GRS Advanced Opportunity Fellowship gennem University of Wisconsin-Madison Graduate School og Bioteknologi Training Program gennem National Institute of General Medical Sciences af National Institutes of Health (T32GM008349).

1. Fremstilling af den horisontale indfødte polyacrylamidgel 4 . <ol><li> Forberedelse af de nødvendige materialer. <ol><li> Til det vandrette gelapparat skal du bruge en gelkasse (37 cm x 24 cm) med en 27 cm x 21 cm bakke og en kapacitet til to 24 brønde kamme. Denne opsætning giver i alt 48 prøver, der kan analyseres samtidigt. </li><li> Forbered følgende reagenser: 40% Acrylamide-Bis 19: 1, 5x TBE (Tris-Borat-EDTA pH 8,0), TEMED og 10% APS (ammoniumpersulfat). </li></o...

<ol>
	<li>Ryder, S. P., Recht, M. I., Williamson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+analysis+of+protein-RNA+interactions+by+gel+mobility+shift.">Quantitative analysis of protein-RNA interactions by gel mobility shift.</a> Methods Mol Biol. 488, 99-115 (2008).</li><li>Dahlberg, A. E., Dingman, C. W., Peacock, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophoretic+characterization+of+bacterial+polyribosomes+in+agarose-acrylamide+composite+gels.">Electrophoretic characterization of bacterial polyribosomes in agarose-acrylamide composite gels.</a> J Mol Biol. 41 (1), 139-147 (1969).</li><li>Hellman, L. M., Fried, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophoretic+mobility+shift+assay+(EMSA)+for+detecting+protein-nucleic+acid+interactions.">Electrophoretic mobility shift assay (EMSA) for detecting protein-nucleic acid interactions.</a> Nat Protoc. 2 (8), 1849-1861 (2007).</li><li>Pagano, J. M., Farley, B. M., Essien, K. I., Ryder, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=RNA+recognition+by+the+embryonic+cell+fate+determinant+and+germline+totipotency+factor+MEX-3.">RNA recognition by the embryonic cell fate determinant and germline totipotency factor MEX-3.</a> Proc Natl Acad Sci USA. 106 (48), 20252-20257 (2009).</li><li>Royer, C. A., Scarlata, S. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescence+approaches+to+quantifying+biomolecular+interactions.">Fluorescence approaches to quantifying biomolecular interactions.</a> Meth Enzymol. 450, 79-106 (2008).</li><li>Su, C., Wang, F., Ciolek, D., Pan, Y. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophoresis+of+proteins+and+protein-protein+complexes+in+native+polyacrylamide+gels+using+a+horizontal+gel+apparatus.">Electrophoresis of proteins and protein-protein complexes in native polyacrylamide gels using a horizontal gel apparatus.</a> Anal Biochem. 223, 93-98 (1994).</li><li>Buczek, P., Horvath, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thermodynamic+Characterization+of+Binding+Oxytricha+nova+Single+Strand+Telomere+DNA+with+the+Alpha+Protein+N-terminal+Domain.">Thermodynamic Characterization of Binding Oxytricha nova Single Strand Telomere DNA with the Alpha Protein N-terminal Domain.</a> J Mol Biol. 359 (5), 1217-1234 (2006).</li><li>Zhang, Y., Park, S., Blaser, S., Sheets, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determinants+of+RNA+binding+and+translational+repression+by+the+Bicaudal-C+regulatory+protein.">Determinants of RNA binding and translational repression by the Bicaudal-C regulatory protein.</a> J Biol Chem. 289 (11), 7497-7504 (2014).</li><li>Gamberi, C., Lasko, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Bic-C+family+of+developmental+translational+regulators.">The Bic-C family of developmental translational regulators.</a> Comp Funct Genomics. , 141386 (2012).</li><li>Saffman, E. E., 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=Premature+translation+of+oskar+in+oocytes+lacking+the+RNA-binding+protein+bicaudal-C.">Premature translation of oskar in oocytes lacking the RNA-binding protein bicaudal-C.</a> Mol Cell Biol. 18 (8), 4855-4862 (1998).</li><li>Chicoine, J., 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=Bicaudal-C+recruits+CCR4-NOT+deadenylase+to+target+mRNAs+and+regulates+oogenesis,+cytoskeletal+organization,+and+its+own+expression.">Bicaudal-C recruits CCR4-NOT deadenylase to target mRNAs and regulates oogenesis, cytoskeletal organization, and its own expression.</a> Dev Cell. 13 (5), 691-704 (2007).</li><li>Zhang, 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=Bicaudal-C+spatially+controls+translation+of+vertebrate+maternal+mRNAs.">Bicaudal-C spatially controls translation of vertebrate maternal mRNAs.</a> RNA. 19 (11), 1575-1582 (2013).</li><li>Maisonneuve, 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=Bicaudal+C,+a+novel+regulator+of+Dvl+signaling+abutting+RNA-processing+bodies,+controls+cilia+orientation+and+leftward+flow.">Bicaudal C, a novel regulator of Dvl signaling abutting RNA-processing bodies, controls cilia orientation and leftward flow.</a> Development. 136 (17), 3019-3030 (2009).</li><li>Tran, U., 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+RNA-binding+protein+bicaudal+C+regulates+polycystin+2+in+the+kidney+by+antagonizing+miR-17+activity.">The RNA-binding protein bicaudal C regulates polycystin 2 in the kidney by antagonizing miR-17 activity.</a> Development. 137 (7), 1107-1116 (2010).</li><li>Pagano, J. M., Clingman, C. C., Ryder, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+approaches+to+monitor+protein-nucleic+acid+interactions+using+fluorescent+probes.">Quantitative approaches to monitor protein-nucleic acid interactions using fluorescent probes.</a> RNA. 17 (1), 14-20 (2011).</li><li>Foley, T. 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=Platform+to+Enable+the+Pharmacological+Profiling+of+Small+Molecules+in+Gel-Based+Electrophoretic+Mobility+Shift+Assays.">Platform to Enable the Pharmacological Profiling of Small Molecules in Gel-Based Electrophoretic Mobility Shift Assays.</a> J Biomol Screen. 21 (10), 1125-1131 (2016).</li></ol>

Indfødte polyacrylamidgeler er et uvurderligt redskab til undersøgelse af protein-RNA-interaktioner, og traditionelt er disse geler elektroforeseret vertikalt 2 , 3 . Vi har anvendt en modifikation af protokollen, som erstatter native polyacrylamidgeler, der er oprettet og elektroforeseret vandret 1 , 4 , 6 , 7 , <sup clas...

Elektroforetiske mobilitetsforskydningsassays (EMSA&#39;er) har vist sig at være et uvurderligt biokemisk værktøj til analyse af specifikke protein-nucleinsyre-interaktioner 1 , 2 , 3 . Disse analyser kan tilvejebringe vigtig information vedrørende proteinbindings bindingsaffinitet til RNA eller DNA 3 , komponentstøkiometrien for nukleinsyre-protein-komplekser 1 og tilvejebringe vigtige nye indsigter om bindingsspecificiteten af ​​RNA-bindingsproteiner via substratkonkurrenceforsøg 1 . Den traditionelle eksperimentelle opsætning for disse analyser består i at blande renset protein med et radioaktivt mærket RNA-substrat. De resulterende komplekser analyseres derefter med ikke-denaturerende (native) polyacrylamidgeler hældt mellem to glasplader efterfulgt af prøveelektroforese i et lodret apparat 3. Selvom denne tilgang er blevet udtømmende anvendt til at tilvejebringe vigtige indsigter i de biokemiske mekanismer, der ligger til grund for bindingen af ​​proteiner til nukleinsyrer, har den også flere begrænsninger. For eksempel har denne grundlæggende strategi relativt lav gennemstrømning, og det er ikke let at tilpasse til applikationer, der kræver analyse af mange bindende reaktioner parallelt. Derudover er det med det traditionelle vertikale apparat udfordrende at potentielt overvåge komplekser ved flere gange under elektroforese 3 , 4 . Her præsenterer vi en tilpasning af EMSA-analysen, der anvender native polyacrylamidgeler, der støbes i et flatbed apparat, vandret elektroforese og fluorescensmærkede RNA-substrater 4 , 5 , 6 , 7 . Indarbejdelsen af ​​disse relativt enkle modifikationer af den basale straTegy giver nogle stærke fordele. Især gør det vandrette flatbed format let at analysere snesevis af prøver samtidigt 4 . For nogle RNA-proteinkomplekser, såsom dem der dannes mellem Bicaudal-C-proteinet og dets RNA-substratelektroforese i en vandret gel, tilvejebringer også en øget evne til at løse forskellige RNA-proteinkomplekser og diskriminere disse fra ubundet RNA-substrat.

<table border="0" cellpadding="0" cellspacing="0" width="1432">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:373px;">Horizontal Gel Box</td>
			<td style="width:112px;">OWL</td>
			<td style="width:116px;">N/A</td>
			<td style="width:831px;">Product no longer made. Similar gel boxes can be found at Thermo Scientific, A-series gel boxes. Catalog Number: A2-BP</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">24-well large large horizontal gel electrophoresis combs</td>
			<td>OWL</td>
			<td>N/A</td>
			<td>Product no longer made. Similar gel boxes can be found at Thermo Scientific, A-series gel box combs. Catalog Number: A2-24C</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Powerpac 300</td>
			<td>Bio-Rad</td>
			<td>1655050</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mini-Protean II Electrophoresis Cell</td>
			<td>Bio-Rad</td>
			<td>165-2940</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">InstaPAGE-19 40% 19:1 Acrylamide/Bis</td>
			<td>IBI</td>
			<td>IB70015</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">TEMED</td>
			<td>IBI</td>
			<td>IB70120</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">APS</td>
			<td>IBI</td>
			<td>IB70080</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Yeast tRNAs</td>
			<td>Ambion</td>
			<td>AM7119</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Fluorescein labeled RNA</td>
			<td>IDT</td>
			<td>N/A</td>
			<td>Order can be made custom to length and desired sequence</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">EDTA tetrasodium salt hydrate</td>
			<td>Sigma-Aldrich</td>
			<td>E5391-1KG</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HEPES</td>
			<td>Sigma-Aldrich</td>
			<td>H4034-500G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tris Base Ultrapure</td>
			<td>US Biological</td>
			<td>T8600</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Boric Acid</td>
			<td>Fisher Scientific</td>
			<td>BP168-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Potassium Chloride</td>
			<td>Fisher Scientific</td>
			<td>BP366-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tween-20</td>
			<td>Fisher Scientific</td>
			<td>BP337-500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">DEPC</td>
			<td>Sigma-Aldrich</td>
			<td>1609-47-8</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Dithiothreitol (DTT)</td>
			<td>Sigma-Aldrich</td>
			<td>3483 12 3</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bovine Serum Albumin (BSA)</td>
			<td>Sigma-Aldrich</td>
			<td>9048-46-8</td>
			<td></td>
		</tr>
	</tbody>
</table>

horizontal gel electrophoresis for enhanced detection of protein-rna complexes

Native polyacrylamidgelelektroforese er et grundlæggende værktøj for molekylærbiologi, der er blevet anvendt i vid udstrækning til den biokemiske analyse af RNA-proteininteraktioner. Disse interaktioner er traditionelt analyseret med polyacrylamidgeler frembragt mellem to glasplader og prøver elektroforeseret vertikalt. Polyacrylamidgeler støbes i bakker og elektroforeses horisontalt, men giver flere fordele. For eksempel kan vandrette geler, som anvendes til analyse af komplekser mellem fluorescerende RNA-substrater og specifikke proteiner, afbildes flere gange, idet elektroforese udvikler sig. Dette giver den unikke mulighed for at overvåge RNA-proteinkomplekser på flere punkter under eksperimentet. Derudover gør horisontal gelelektroforese det muligt at analysere mange prøver parallelt. Dette kan i høj grad lette tidskursforsøg samt analysere flere reaktioner samtidigt for at sammenligne forskellige komponenter og betingelser. Her prOvide en detaljeret protokol til generering og anvendelse af horisontal indfødt gelelektroforese til analyse af RNA-Protein-interaktioner.

For at demonstrere kraften og alsidigheden af ​​den horisontale indfødte gelelektroforese analyserede vi bindingen af Xenopus Bicaudal-C (Bicc1) -proteinet til et fluorescensmærket RNA indeholdende et Bicc1-bindingssted. Bicc1-proteiner virker som mRNA-specifikke translationsundertrykkere for at kontrollere beslutninger om cellernes skæbne under moderfasen af ​​dyreudvikling 9 , 10 , <sup class="xre...

Watch this Scientific Journal Video about Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes at JoVE.com

Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes

Native polyacrylamidgelelektroforese er et grundlæggende redskab til analyse af RNA-proteininteraktioner. Traditionelt har de fleste eksperimenter brugt vertikale geler. Imidlertid tilvejebringer vandrette geler flere fordele, såsom muligheden for at overvåge komplekser under elektroforese. Vi leverer en detaljeret protokol til generering og anvendelse af horisontal indfødte gelelektroforese.

Horisontal gelelektroforese til forbedret påvisning af protein-RNA-komplekser

Native polyacrylamide gel electrophoresis is a fundamental tool of molecular biology that has been used extensively for the biochemical analysis of ...

The overall of this protocol is to analyze RNA protein interactions using horizontal native gel electrophoresis. This method can help answer key questions in RNA biochemistry, such as defining the characteristics of specific RNA protein interactions. The main advantage of this technique is that it allows complex formation to be monitored multiple times during electrophoresis.For the horizontal gel apparatus, use a gel box with a 27 centimeter by 21 centimeter casting tray and a capacity for two 24 well combs. This set up provides a total of 48 samples that can be analyzed simultaneously. Prepare two liters of 1x TBE running buffer by diluting 400 milliliters of 5x TBE with 1, 600 milliliters of water and mixing.Place the TBE buffer in the cold room to cool. Prepare the following reagents following standard procedures, 40%acrylamide best 19:1, 5x TBE, temed and 10%ammonium per sulfate. To make a 10%native acrylamide gel, add the following to a 500 milliliter flask:80 milliliters of 5x TBE, 100 milliliters of 40%acrylamide and water to a final volume of 400 milliliters.Add four milliliters of 10%ammonium per sulfate and 400 microliters of temed and mix well. Pour the mix into the horizontal casting tray and allow the gel to polymerize in the fume hood for 30 minutes. The polymerized gel will be approximately two centimeters thick and a thin layer of liquid will remain on top of the gel.Place the gel into the gel box and take it into the cold room. Next, pour the liquid off and rinse with running buffer. Remove the combs carefully, add two liters of cold running buffer to the gel box and then use a syringe to rinse the wells with running buffer.Pre-run the gel at 120 volts for one hour at four degrees celsius in a cold room. While the gel is being prerun, prepare the binding reactions. Add the following binding reaction components to each RNA free microcentrifuge tube in this order:five microliters of 10 millimolar BSA, five microliters of 20 millimolar DTT, five microliters of 10 millimolar yeast tRNAs, 10 microliters of 5x binding buffer, bicaudal-c protein to a final concentration of 250 nanomolar in 50 microliters, DEPC-treated water to a final volume of 45 microliters and five microliters of fluorescently labeled RNA substrate.Close the tubes and flick gently to mix, centrifuge the tubes briefly and incubate in the dark for 30 minutes at room temperature. When the binding reactions are complete, add 15 microliters of 50%glycerol to each reaction and mix gently. Carefully load 30 microliters of each reaction into the wells of the gel, the amount of sample that can be loaded into a single well varies depending on the thickness of the gel and the size of the wells.The power supply to the gel apparatus, switch on the power, adjust the voltage to 120 volts and run the gel in four degree celsius. To prevent damaging or bleaching of the fluorescently labeled RNA, conduct electrophoresis in the dark. Electrophoresis times will vary depending upon the specifics of the RNA protein complex being analyzed.A major advantage of the horizontal electrophorated mobility shift assay analysis, is that electrophoresis can be stopped in the gel image prior to completion of the run. Perform analysis with any instrument designed for detecting an imaging fluorescent substrates. Adjust the imager settings, for fluorescently labeled ligands, use the following settings.Fluorescein wavelength of 473 nanometers, photomultiplier voltage of 750 volts and pixel size of 100 micrometers. Scan the gel to capture the image. After imaging, the gel can be placed back into the apparatus and electrophoresis can be continued for longer times.The binding of the xenopus bicaudal-C or Bicc1 protein to a fluorescently labeled Cripto1 mRNA containing a Bicc1 binding site, was analyzed by both vertical and horizontal native gel electrophoresis, as a negative control, Bicc1 was mixed with a fluorescently labeled RNA derived from CyclinB1 mRNA. It was readily apparent using either method, that Bicc1 binds to the Cripto1 RNA and forms a specific complex. However, the Bicc1, Cripto1 complexes were significantly more distinct when analyzed with the horizontal gel, facilitating their detection and allowing the discrimination of protein RNA complexes from unbound RNA.After imaging, the gel was returned to the gel apparatus, electrophoresis for an additional three and a half hours, and reimaged. As shown here, the complexes had migrated further and were still readily detectable, indicating their stability. Once mastered, this technique can be done in four to five hours, if it is performed properly.After watching this video, you should have a good understanding of how to analyze RNA protein interactions using horizontal native gel electrophoresis.

Research

JoVE Journal

Biochemistry

Sheathless Capillary Electrophoresis&#8211;Mass Spectrometry for Metabolic Profiling of Biological Samples

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In metabolomics, a wide range of analytical techniques is used for the global profiling of (endogenous) metabolites in complex samples. In this paper, a ...

Characterization of Multi-subunit Protein Complexes of Human MxA Using Non-denaturing Polyacrylamide Gel-electrophoresis

Karakterisering af Multi-subunit proteinkomplekser af menneskelig MxA Brug ikke-denaturering Polyacrylamid Gel-elektroforese

The formation of oligomeric complexes is a crucial prerequisite for the proper structure and function of many proteins. The interferon-induced antiviral ...

Tandem Affinity Purification of Protein Complexes from Eukaryotic Cells

Tandem affinitetsoprensning af proteinkomplekser fra eukaryote celler

The purification of active protein-protein and protein-nucleic acid complexes is crucial for the characterization of enzymatic activities and de novo ...

Protein-tRNA Agarose Gel Retardation Assays for the Analysis of the N6-threonylcarbamoyladenosine TcdA Function

Protein-tRNA Agarose Gel Retardation Assays for the Analysis of the N6-threonylcarbamoyladenosine TcdA Function

Protein-tRNA Agarose Gel Retardation Assays for Analysen af<em&gt; N</em<sup&gt; 6</sup&gt; -threonylcarbamoyladenosin TcdA-funktion

We demonstrate methods for the expression and purification of tRNA(UUU) in Escherichia coli and the analysis by gel retardation assays of the binding of ...

Detection and Visualization of DNA Damage-induced Protein Complexes in Suspension Cell Cultures Using the Proximity Ligation Assay

Påvisning og visualisering af DNA-skader-inducerede proteinkomplekser i suspensionscellekulturer ved anvendelse af Proximity Ligation Assay

The DNA damage response orchestrates the repair of DNA lesions that occur spontaneously, are caused by genotoxic stress, or appear in the context of ...

Analyzing Supercomplexes of the Mitochondrial Electron Transport Chain with Native Electrophoresis, In-gel Assays, and Electroelution

Analyse af superkomplekser af den mitokondrieelektron transportkæde med indfødte elektroforese, in-gel analyser og elektrolysering

The mitochondrial electron transport chain (ETC) transduces the energy derived from the breakdown of various fuels into the bioenergetic currency of the ...

DNA Polymerase Activity Assay Using Near-infrared Fluorescent Labeled DNA Visualized by Acrylamide Gel Electrophoresis

DNA Polymerase aktivitet Assay ved hjælp af nær-infrarødt fluorescerende mærket DNA visualiseret af acrylamid Gel elektroforese

For any enzyme, robust, quantitative methods are required for characterization of both native and engineered enzymes. For DNA polymerases, DNA synthesis ...

Use of Two Dimensional Semi-denaturing Detergent Agarose Gel Electrophoresis to Confirm Size Heterogeneity of Amyloid or Amyloid-like Fibers

Brugen af to-dimensionelle semi-denaturering vaskemiddel Agarosen gelelektroforese at bekræfte størrelse heterogenitet af Amyloid eller Amyloid-lignende fibre

Amyloid or amyloid-like fibers have been associated with many human diseases, and are now being discovered to be important for many signaling pathways. ...

Analysis of AtHIRD11 Intrinsic Disorder and Binding Towards Metal Ions by Capillary Gel Electrophoresis and Affinity Capillary Electrophoresis

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Plants are strongly dependent on their environment. In order to adjust to stressful changes (e.g., drought and high salinity), higher plants evolve ...

Analysis of Thylakoid Membrane Protein Complexes by Blue Native Gel Electrophoresis

Analyse af tylakoid membran proteinkomplekser af blå indfødte gelelektroforese

Photosynthetic electron transfer chain (ETC) converts solar energy to chemical energy in the form of NADPH and ATP. Four large protein complexes embedded ...