Name: generation of transgenic c. elegans by biolistic transformation
Uploaded: 2010-08-23T17:00:00
Description: Watch this Scientific Journal Video about Generation of Transgenic C. elegans by Biolistic Transformation at JoVE.com

Questo lavoro è stato sostenuto da fondi di avviamento presso l&#39;Università di Pittsburgh e NIH concedere AG028977 a ALF

Panoramica Tradizionalmente vermi transgenici sono stati generati da microinjecting DNA transgene nel C. elegans germinale 1,2,3,4. Anche se di successo, questo approccio richiede attrezzature specializzate e lo sviluppo di esperienza con l&#39;anatomia vite senza fine e la tecnica di microiniezione. Più di recente bombardamento biolistic è stato sviluppato come un approccio alternativo che utilizza rivestite di DNA particelle d&#39;oro per introdurre DNA estraneo nella l...

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Bombardamento Biolistic è un metodo semplice per introdurre DNA estraneo in molti organismi, tra cui C. elegans 1,5,6,7,16. Si basa su particelle d&#39;oro che formano un complesso con il DNA in presenza di CaCl 2. Poliammine cationiche, come spermidina, proteggere il DNA dalla degradazione nucleasi in vivo. Dal momento che spermidina è una molecola labile, è importante conservarlo in piccole aliquote a -20 ° C, e fare la giusta soluzione prima di eseguire il bombardamento. Come abbiam...

The volume for a solution in, <a href="http://www.jove.com/video/2090">Generation of Transgenic C. elegans by Biolistic Transformation</a>, was incorrect.

Erratum: Generation of Transgenic C. elegans by Biolistic Transformation

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<td>Gold particles</td>
<td>&nbsp;</td>
<td>Inbio gold</td>
<td>BD021</td>
<td>1micron</td>
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<td>Midiprep kit</td>
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<td>Qiagen</td>
<td>12143</td>
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<td>&nbsp;</td>
<td>Sigma</td>
<td>S4139</td>
<td>&nbsp;</td>
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<td>Protamine</td>
<td>&nbsp;</td>
<td>Sigma </td>
<td> P4505</td>
<td>&nbsp;</td>
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<td>Macrocarriers</td>
<td>&nbsp;</td>
<td>Biorad</td>
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generation of transgenic c. elegans by biolistic transformation

Il numero di laboratori che si servono libero nematode C. vita elegans è in rapida crescita. La popolarità di questo modello biologico è attribuita ad un tempo di generazione rapida e breve vita, la manutenzione facile e poco costoso, genoma completamente sequenziato, e vasta gamma di risorse RNAi e animali mutanti. Inoltre, l&#39;analisi della C. elegans genoma ha rivelato una grande somiglianza tra vermi e vertebrati superiori, il che suggerisce che la ricerca di vermi potrebbe essere un&#39;aggiunta importante a studi condotti nei topi tutto o in cellule in coltura. Un aspetto importante della ricerca e worm è la capacità di utilizzare animali transgenici per studiare la localizzazione dei geni e la funzione. Animali transgenici possono essere create sia tramite microiniezione della linea germinale verme o attraverso l&#39;uso di bombardamento biolistic. Bombardamento è una tecnica più nuova ed è meno familiare ad una serie di laboratori. Qui descriviamo un semplice protocollo per generare i vermi transgenici dai bombardamenti biolistic con particelle d&#39;oro con il Bio-Rad sistema PDS-1000. Rispetto a microiniezione del DNA in ermafrodita germinale, questo protocollo ha il vantaggio di non richiedere particolari competenze da parte dell&#39;operatore per quanto riguarda l&#39;identificazione o l&#39;esecuzione di anatomia verme microiniezione. Ulteriori più linee transgeniche sono solitamente ottenuti da un singolo bombardamento. Anche in contrasto con microiniezione, bombardamento biolistic produce animali transgenici con entrambi gli array extracromosomico e transgeni integrato. La possibilità di ottenere integrato linee transgeniche possono evitare l&#39;uso di protocolli mutageno di integrare DNA estraneo. In conclusione, bombardamento biolistic può essere un metodo interessante per la generazione di animali transgenici, soprattutto per gli investigatori non sono interessati a investire il tempo e lo sforzo necessario per diventare abili a microiniezione.

Watch this Scientific Journal Video about Generation of Transgenic C. elegans by Biolistic Transformation at JoVE.com

Generation of Transgenic C. elegans by Biolistic Transformation

Vermi transgenici sono comunemente utilizzati in<em&gt; C. elegans</em&gt; Ricerca. Descritto è un protocollo semplice, ma efficace, di introdurre transgeni in vermi mediante bombardamento biolistic con DNA particelle rivestite d&#39;oro. Lo sforzo ei risultati del bombardamento reggono bene il confronto con microiniezione per la produzione di animali transgenici.

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The number of laboratories using the free living nematode C. elegans is rapidly growing. The popularity of this biological model is attributed to a rapid ...

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Generation of Transgenic C. elegans by Biolistic Transformation

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Transgenic Caenorhabditis elegans can be readily created via microinjection of a DNA plasmid solution into the gonad 1.  The plasmid DNA rearranges to ...

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In this protocol, we present the required materials, and the procedure for making modified&#160;C. elegans&#160;Habituation and Reproduction media ...