Difference between revisions of "Team:CPU CHINA/Experiments"

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var zhuanran = "<h2>Cell transfection and Screening</h2>"+
 
var zhuanran = "<h2>Cell transfection and Screening</h2>"+
 
"<h3>1.Cell transfection</h3>"+
 
"<h3>1.Cell transfection</h3>"+
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"<h4>HepG2 cells were cultured in DEME medium containing 10% heat inactivated bovine serum at 37 ℃ and 5% CO2 constant temperature incubator. HepG2 cells were seeded in 96-well plate according to 2*10<sup>5</sup>/ well, and cultured in non-antibiotic culture medium for 24 hours, then transfected with liposome (<a><u>Lipofectamine 3000</u></a>) , and transfected with 3 holes per plasmid. The transfection medium was replaced after 4 to 6 h, and the DEME culture medium containing 10% calf serum was used to continue the culture. </h4><br>"+
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"<h4>(1) Culture HepG2 cells with 10% heat inactivated bovine serum (Gibco, US) in DEME medium  and a 5% CO2 constant temperature incubator at 37 ℃.</h4><br>"+
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                "<h4>(2) Seed HepG2 cells in a 96-well plate at a concentration of 2*10<sup>5</sup>/well, and culture them in non-antibiotic culture medium (Gibco, US) for 24 hours. </h4><br>"+
 +
"<h4>(3) Transfect the cell with Lipofectamine 3000 (Thermofisher, US).</h4><br>"+
 +
                "<h4>(4) Refresh the medium at 4h to 6h after the transfection with DEME culture medium containing 10% calf serum (Gibco, US). </h4><br>"+
 
"<h3>2.Screening</h3>"+
 
"<h3>2.Screening</h3>"+
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"<h4>After 24-48 h, the DEME culture medium containing antibiotics (Neomycin) was used to screen.</h4><br>"+
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"<h4>After 24-48 h, use the DEME culture medium containing antibiotics (Neomycin) (Gibco, US) for the screening of cells successfully transfected with plasmids. </h4><br>"+
 
 
 
"<h2>Luciferase reporter gene assay</h2>"+
 
"<h2>Luciferase reporter gene assay</h2>"+
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"<h4>pGL3-Basic was used as negative Control, and pGL3-Control was used as positive Control. HepG2 cells were transfected with 1ug of pGL3-htert, pGL3-hulc and pGL3-control(Refer Cell transfection and Screening-Part.1).<h4>"+
 
 
"<h4>After 48 h, the cells were treated with Promege's <a><u>steady-glo Luciferase Assay System kit </u></a>and the activity of <i>Luc</i> was determined.</h4><br>";
 
"<h4>After 48 h, the cells were treated with Promege's <a><u>steady-glo Luciferase Assay System kit </u></a>and the activity of <i>Luc</i> was determined.</h4><br>";
 
 

Revision as of 10:06, 7 December 2018

Experiments

Molecular Cloning

Transfection

You can click on the figure to see the protocol.

Characterization

You can click on these arrows to read the details of our experiments.

You can click here to see our Promoter.