Haynes Lab:Notebook/RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines/2014/12/16: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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==Summary== | ==Summary== | ||
In order to introduce a new gene into the mammalian cancer cells, we need to make the cells competent and present them with a prepared plasmid containing our gene of interest along with a selectable marker. Lipofectamine is a tranfection reagent that produces positively charged liposomes that envelop the plasmids to be introduced to the cells and encourage passage through the negatively charged cell membrane into the cells. | |||
==Goals== | ==Goals== | ||
* | * Begin transfection of U2OS cells in 6-well plate<br><br> | ||
==Protocol== | ==Protocol== | ||
[http://openwetware.org/wiki/Haynes: | [http://openwetware.org/wiki/Haynes:TransfectionPlasmid_Lipo mammalian cell transfection with liopfectamine]<br><br> | ||
==Materials== | ==Materials== | ||
* | * Lipofectamine LTX | ||
* | * Opti-MEM I Reduced serum medium | ||
* Antibiotic-free growth medium (no pen/strep or other antibiotics) | |||
* Cells | |||
* Pipette tips<br><br> | |||
==Notes== | ==Notes== | ||
To make the transfection solution, in a 1.5mL microcentrifuge tube add: | |||
* 20μL DNA in H<sub>2</sub>O | |||
* 570μL Opti-MEM solution | |||
* 2.5μL PLUS reagent | |||
* wait 5 minutes at room temp. | |||
* 7.5μL Lipofectamine LTX | |||
* wait 30 minutes at room temp. | |||
* begin transfection<br><br> | |||
==Results/Conclusions== | ==Results/Conclusions== | ||
Three wells were treated with DNA, one was treated with water (negative control). Well plate is in the upper incubator, middle shelf.<br><br> | |||
*'''[[User:David Benjamin Nyer|David Benjamin Nyer]] 18: | *'''[[User:David Benjamin Nyer|David Benjamin Nyer]] 18:36, 16 December 2014 (EST)''': | ||
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__NOTOC__ | __NOTOC__ |
Latest revision as of 00:36, 27 September 2017
RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines | Main project page Previous entry Next entry |
SummaryIn order to introduce a new gene into the mammalian cancer cells, we need to make the cells competent and present them with a prepared plasmid containing our gene of interest along with a selectable marker. Lipofectamine is a tranfection reagent that produces positively charged liposomes that envelop the plasmids to be introduced to the cells and encourage passage through the negatively charged cell membrane into the cells. Goals
Protocolmammalian cell transfection with liopfectamine Materials
NotesTo make the transfection solution, in a 1.5mL microcentrifuge tube add:
Results/ConclusionsThree wells were treated with DNA, one was treated with water (negative control). Well plate is in the upper incubator, middle shelf.
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