Name: rnai-mediated gene knockdown and in vivo diuresis assay in adult female aedes aegypti mosquitoes
Uploaded: 2012-07-14T14:00:00
Description: Watch this Scientific Journal Video about RNAi-mediated Gene Knockdown and In Vivo Diuresis Assay in Adult Female Aedes aegypti Mosquitoes at JoVE.com

De auteurs danken Victoria Carpenter voor haar kritische opmerkingen van dit protocol.

Deel I - RNAi-gemedieerde gen knock-down in het volwassenenonderwijs Aedes aegypti muggen. Voor experiment overzicht zie figuur 1. 1. dsRNA Synthese <ol><li> Synthetiseren specifieke dsRNA&#39;s tegen genen van belang en controle dsRNA&#39;s. Opmerking: we raden ontwikkeling primers voor PCR fragmenten in het bereik van 300 tot 500 baseparen gelegen aan het 3 &#39;uiteinde van de specifieke cDNA 2 en het T7 primersequentie bevestigd aan het 5&#39;-uiteinde...

<ol>
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De gebruikte RNAi-protocol is ontwikkeld in het laboratorium van Alexander Raikhel aan de Universiteit van Californië Riverside 6,7 en is vergelijkbaar met een protocol gepubliceerd door Garver en Dimopoulos 4. De experimentele benadering in deze video protocol kan worden gebruikt om genen in insect diurèse in een in vivo studie omgeving. De uitscheidingsorganen van insecten, de buisjes van Malpighi, hebben de interesse van de generaties van onderzoekers als een &#39;simpele&#39; modelsy...

<table border="1"><tbody><tr><td> Naam van het reagens of apparatuur </td><td> Vennootschap </td><td> Catalogusnummer </td><td> Reacties </td></tr><tr><td> MEGAscript T7 High Yield Kit </td><td> Ambion, Inc </td><td> AM1334 </td><td></td></tr><tr><td> PBS buffer </td><td> Sigma-Aldrich </td><td> P4417 </td><td></td></tr><tr><td> Kunststof leidingen </td><td> Lokale leverancier </td><td></td><td> PVC </td></tr><tr><td> 1 ml kunststof pipetpunt </td><td> VWR </td><td> 83007-376 </td><td> Blue tip </td></tr><tr><td> 1 ml spuit </td><td> Becton, Dickinson and Company </td><td> 309602 </td><td></td></tr><tr><td> Schaar </td><td> Lokale leverancier </td><td></td><td></td></tr><tr><td> Metalen naald </td><td> Carolina Biologicals </td><td> 654307 </td&gt; <td> Maat 5 </td></tr><tr><td> Fly pad </td><td> Genesee Wetenschappelijk </td><td> 789060 </td><td></td></tr><tr><td> Batterij-aangedreven aspirator w / collectie flacon </td><td> Upma Labs </td><td> IPMM 2000 </td><td></td></tr><tr><td> Fijne punt tang </td><td> Wereld Precision Instruments </td><td> 14095 </td><td></td></tr><tr><td> Glascapillair naalden </td><td> Wereld Precision Instruments </td><td> 1B200-6 </td><td></td></tr><tr><td> Stereo dissectie microscoop </td><td> Leica Microsystems </td><td> S6D </td><td></td></tr><tr><td> Analytische precisiebalans </td><td> Mettler Toledo </td><td> AB54S </td><td></td></tr><tr><td> Sucrose </td><td> Sigma-Aldrich </td><td> 84097 </td><td></td></tr><tr><td> Een pint was gezet gevoerd kartonnen bekers </td><td> Lokale leverancier </td><td></td><td> Gefabriceerd soep bekers </td></tr><tr><td> Mesh netto </td><td> Lokale leverancier </td><td></td><td> plastic vliegen gaas </td></tr></tbody></table>

rnai-mediated gene knockdown and in vivo diuresis assay in adult female aedes aegypti mosquitoes

Deze video protocol geeft blijk van een effectieve techniek om een ​​bepaald gen in een insect knockdown en voert een nieuwe bioassay om de uitscheiding te meten. Deze methode kan worden gebruikt om een ​​beter begrip van de werkwijze volgens diurèse in insecten verkrijgen en is bijzonder nuttig bij het bestuderen van diurèse in bloed geeft geleedpotigen die kunnen nemen grote hoeveelheden vloeistof in een bloedmaaltijd. Deze RNAi-gemedieerde gen-knockdown in combinatie met een in vivo diurese test werd ontwikkeld door de Hansen lab om de effecten van RNAi-gemedieerde knockdown van aquaporine genen op Aedes aegypti mug diurese 1 te bestuderen. Het protocol is ingesteld in twee delen: de eerste demonstratie laat zien hoe een eenvoudige injectie mug apparaat te maken en hoe voor te bereiden en dsRNA te injecteren in de thorax van muggen voor RNAi-gemedieerde gen knockdown. De tweede demonstratie illustreert hoe bepalenuitscheiding prijzen voor muggen met behulp van een in vivo bioassay.

Watch this Scientific Journal Video about RNAi-mediated Gene Knockdown and In Vivo Diuresis Assay in Adult Female Aedes aegypti Mosquitoes at JoVE.com

RNAi-mediated Gene Knockdown and In Vivo Diuresis Assay in Adult Female Aedes aegypti Mosquitoes

In dit protocol combineren we RNAi-gemedieerde gensilencing een<em&gt; In vivo</em&gt; Diurese test om de effecten knockdown van genen van belang te bestuderen heeft op muggen vloeistof excretie.

RNAi-gemedieerde Gene Knockdown en<em&gt; In Vivo</em&gt; Urineproductie Assay in het volwassenenonderwijs Vrouw<em&gt; Aedes aegypti</em&gt; Muggen

This video protocol demonstrates an effective technique to knockdown a particular gene in an insect and conduct a novel bioassay to measure excretion ...

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RNAi-mediated Gene Knockdown and In Vivo Diuresis Assay in Adult Female Aedes aegypti Mosquitoes

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Mosquitoes represent a significant public health problem as vectors of various pathogens. For those studies that require an assessment of mosquito fitness ...