Name: the production of c. elegans transgenes via recombineering with the galk selectable marker
Uploaded: 2011-01-11T23:58:00
Description: Watch this Scientific Journal Video about The Production of C. elegans Transgenes via Recombineering with the galK Selectable Marker at JoVE.com

저자는 기술을 개발에 대한 도움 린지 내쉬 감사하고 싶습니다. 이 작품은 ALF, 피츠버그 대학에서 피츠버그 OAIC 대학 (AG024827) 및 씨앗 기금에서 파일럿 프로젝트 부여하는 NIH 부여 AG028977에 의해 투자되었다.

개요 형질 C.의 생성에 사용되는 많은 transgenes 엘레간스는 모터의 시퀀스로 구성 아마도 cDNA 유전자 박사 앤디 화재 1의 실험실에 의해 생성된 벡터 중 하나에 복제. 이러한 transgenes 자주 GFP의 리포터 유전자를 생산하거나 원하는 패턴의 cDNA를 표현 관련하여 성공하는 동안 이러한 transgenes는 다른 발기인, 확장기 요소 및 컨트롤에서 중요한 역할을 3 &#39;번?...

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fosmids에서 transgenes의 생성은 기본 발기인 요소, 스플 라이스 변종, 3 &#39;UTR 규제 요소를 모두 유지의 혜택을 제공합니다. 이것은 기본 표현식 패턴, 또는 다른 접근법 5 실패 기능성 transgene의 건설이 더 반영하는 transgene의 건설로 이어질 수 있습니다. 결과 transgenes는 GFP 또는 TAP 태그를 포함한 에피토프 태그의 다양한 휴대하실 수 있습니다. transgenes의 건설은 모든 SW106...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>FosmidMAX kit</td>
<td>&nbsp;</td>
<td>Epicentre</td>
<td>FMAX046</td>
<td>&nbsp;</td>
<tr>
<td>GoTaq</td>
<td>&nbsp;</td>
<td>Promega</td>
<td>M7122</td>
<td>&nbsp;</td>
<tr>
<td>MOPS Media</td>
<td>&nbsp;</td>
<td>Teknova</td>
<td>M2120</td>
<td>&nbsp;</td>
<tr>
<td>0.132 M Potassium phosphate solution</td>
<td>&nbsp;</td>
<td>Teknova</td>
<td>M2102</td>
<td>&nbsp;</td>
<tr>
<td>D-galactose</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>G0750</td>
<td>&nbsp;</td>
<tr>
<td>2-deoxygalactose</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>D4407</td>
<td>&nbsp;</td>
<tr>
<td>Biotin</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>B4639</td>
<td>&nbsp;</td>
<tr>
<td>Leucine</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>L8000</td>
<td>&nbsp;</td>
<tr>
<td>NH4Cl</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>A9434</td>
<td>&nbsp;</td>
<tr>
<td>Phusion DNA polymerase</td>
<td>&nbsp;</td>
<td>NEB</td>
<td>F-530S</td>
<td>&nbsp;</td>
<tr>
<td>MacConkey agar base</td>
<td>&nbsp;</td>
<td>Becton Dickinson</td>
<td>281810</td>
<td>&nbsp;</td>
<tr>
<td>Arabinose</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>A3131</td>
<td>&nbsp;</td>
<tr>
<td>Chloramphenicol</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>C1919</td>
<td>&nbsp;</td>
<tr>
<td>Sodium phosphate dibasic</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>S5136</td>
<td>&nbsp;</td>
<tr>
<td>Potassium phosphate monobasic</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P5655</td>
<td>&nbsp;</td>
<tr>
<td>Sodium chloride</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>S5886</td>
<td>&nbsp;</td>
<tr>
<td>Glycerol</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>G2025</td>
<td>&nbsp;</td>
<tr>
<td>Bacto Agar</td>
<td>&nbsp;</td>
<td>Becton Dickinson</td>
<td>214010</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

the production of c. elegans transgenes via recombineering with the galk selectable marker

유전자 변형 동물의 생성은 널리 C로 활용 규정과이자 또는 탠덤 친화 정화의 세대 (TAP) 특정 유전자의 버전이 자신의 정화를 촉진하는 태그의 유전자의 표현 패턴을 연구하기 위해 GFP 융합 단백질의 사용을 포함 엘레간스 연구. 일반적으로 transgenes는 GFP의 리포터 유전자 또는 cDNA의 관심의 상류 발기인을 배치에 의해 생성되며, 이것은 종종 대표 표현식 패턴을 생산하고 있습니다. 그러나, 3 &#39;번역되지 않은 지역이나 대안 발기인의 제어 요소와 같은 유전자 조절의 중요한 요소는,이 접근법에 의해보고 싶었어 수 있습니다. 또한 단 하나의 스플 라이스 변종은 일반적으로이 방법으로 공부하실 수 있습니다. 반면, fosmid의 DNA를 복제하여 실시 웜 게놈의 DNA의 사용 가능성이 포함 대부분의 진정한 표현의 패턴과 타이밍을 캡처하는 더 큰 능력을 허용하는 생체내의 유전자 조절에 관련된 모든 요소하지 않을 경우. fosmid DNA를 사용하여 transgenes의 생성을 촉진하기 위해, 우리는 E.을 설명 대장균의 유전자에 위치에 GFP, TAP 태그 또는 관심의 다른 시퀀스를 삽입하는 recombineering 절차를 기반으로. 절차는 높은 효율로 원하는 수정을 얻는 결과 recombineering에 긍정적이고 부정적인 선택 단계 모두 선택 마커로 galK 유전자를 사용합니다. 또한, 일반적으로 사용되는 GFP에 상동의 무기 둘러싸인과 융합 유전자를 TAP galK 유전자를 포함하는 plasmids는 GFP 또는 탭 융합 단백질을 생성할 때 50 % oligos의 비용을 줄일 수있는 사용할 수 있습니다. 이러한 plasmids은 광범위한 PCR 제품 정화의 필요성을 걸로 R6K 복제 원점을 사용합니다. 마지막으로, 우리는 또한 fosmid 직접 주입 또는 유전자 변형 동물을 생성하는 벌레로 폭격을 할 수있는 fosmid 백본에 UNC - 119 마커를 통합하는 방법을 보여줍니다. 이 동영상은이 방법을 사용 recombineering 통해 transgene을 생성에 관련된 절차를 보여줍니다.

Watch this Scientific Journal Video about The Production of C. elegans Transgenes via Recombineering with the galK Selectable Marker at JoVE.com

The Production of C. elegans Transgenes via Recombineering with the galK Selectable Marker

대한 transgenes를 생산하는 능력<em&gt; Caenorhabditis 엘레간스</em&gt; 기본 규제 요소가 모두 유지됩니다로서 fosmids에 의해 실시 게놈 DNA를 사용하는 것이 특히 매력적이다. transgenes의 생산을위한 간단하고 강력한 절차와 recombineering 통해이다 설명<em&gt; galK</em&gt; 선택 마커.

생산 C. 엘레간스</em함께 Recombineering 통해> Transgenes galK 선택 마커

The creation of transgenic animals is widely utilized in C. elegans research including the use of GFP fusion proteins to study the regulation and ...

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The Production of C. elegans Transgenes via Recombineering with the galK Selectable Marker

Generation of Stable Transgenic C. elegans Using Microinjection

Microinjection을 사용하여 안정적인 트렌스​​ 제닉 C. 엘레간스의 생성

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생산 C. 엘레간스 Transgenes galK 선택 마커