Difference between revisions of "Team:ICT-Mumbai/Protocols"

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<button class="accordion">Preparation of <i>E. coli</i> competent cells</button>
 
<button class="accordion">Preparation of <i>E. coli</i> competent cells</button>
 
<div class="panel">
 
<div class="panel">
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   <p> <i>Materials</i>
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   <p> <i>Materials</i> <br>
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1. Overnight E. coli culture
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1. Overnight E. coli culture<br>
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2. 10 ml LB medium in culture tubes
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2. 10 ml LB medium in culture tubes<br>
 
3. 0.1 M CaCl2
 
3. 0.1 M CaCl2
 
4. 0.1 M CaCl2 containing 25% glycerol
 
4. 0.1 M CaCl2 containing 25% glycerol

Revision as of 15:17, 29 October 2017

ICT-Mumbai 2017

Materials
1. Overnight E. coli culture
2. 10 ml LB medium in culture tubes
3. 0.1 M CaCl2 4. 0.1 M CaCl2 containing 25% glycerol Equipment 1. Spectrophotometer or plate reader 2. Centrifuge Procedure 1. Subculture 100 µl overnight grown culture of E. coli in 10 ml LB medium. Shake at 200 rpm at 37°C. 2. After ~2 hours, using either a spectrophotometer or a plate reader, measure absorbance of the culture at 600 nm. At OD600 = 0.2, harvest the cells by centrifugation. 3. Wash cells with ice cold solution of 0.1 M CaCl2 two to three times. 4. Resuspend cells in 100 μl of 0.1 M CaCl2 containing 25% glycerol and store at –80°C till further use.

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