Team:Elan Vital South Korea/l transformation
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<h1 class="title">Transformation</h1> | <h1 class="title">Transformation</h1> | ||
- | <p class="listing">a. Take out 50㎕of component cell kept in 80 degrees below zero and melt it in ice. (We used the compotent cell of One Shot Top 10 Chemically compotent E. coli from Invitrogen.)</p> | + | <p class="listing">a. Take out 50㎕of component cell<i class="green">*</i> kept in 80 degrees below zero and melt it in ice. (We used the compotent cell of One Shot Top 10 Chemically compotent E. coli from Invitrogen.)</p> |
- | <p class="listing">b. Add 1㎕ of plasmid DNA to each of the tube.</p> | + | <p class="listing">b. Add 1㎕ of plasmid DNA<i class="green">**</i> to each of the tube.</p> |
<p class="listing">c. Mix well and place them in ice for 30 minutes.</p> | <p class="listing">c. Mix well and place them in ice for 30 minutes.</p> | ||
<p class="listing">d. Place plasmid DNA in the compotent cell and keep it in 42℃ for 30 seconds and place it in ice for 2 minutes.</p> | <p class="listing">d. Place plasmid DNA in the compotent cell and keep it in 42℃ for 30 seconds and place it in ice for 2 minutes.</p> | ||
- | <p class="listing">e. Add 250㎕ of liquid LB | + | <p class="listing">e. Add 250㎕ of liquid LB broth in each of the tube and grow it in 37℃ shaking incubator for an hour. </p> |
<p class="listing">f. Leave 100㎕ and dispose.</p> | <p class="listing">f. Leave 100㎕ and dispose.</p> | ||
- | <p class="listing">g. Resuspend the cell, and spread 25㎕ in each of the ampicillin, kanamycin, gentamycin, tetracycline LB solid | + | <p class="listing">g. Resuspend the cell, and spread 25㎕ in each of the ampicillin, kanamycin, gentamycin, tetracycline LB solid plate.</p> |
- | <p class="listing">h. Wait until the E. coli bacteria smears into the plate.</p> | + | <p class="listing">h. Spread 25㎕ of competent E. coli in LB solid plate with the same antibiotics.</p> |
- | <p class="listing"> | + | <p class="listing">i. Wait until the E. coli bacteria smears into the plate.</p> |
+ | <p class="listing">j. Grow it in the 37℃ incubator for a day and observe.</p> | ||
+ | <p class="listing">k. Go through miniprep again.</p> | ||
+ | <p class="listing_sub green">*We used the competent cell from ‘One Shot® TOP10 Chemically competent E. coli’ from Introgen.</p> | ||
+ | <p class="listing_sub green">**We used the plasmid DNA we got from miniprep previously.</p> | ||
+ | <img class="wiki_img" src="https://static.igem.org/mediawiki/2014hs/7/7b/Elan_vital_process_1.png" /> | ||
+ | <p class="wiki_caption">How transformation works</p> | ||
+ | <img class="wiki_img" src="https://static.igem.org/mediawiki/2014hs/6/66/Elan_vital_protocol_1.png" /> | ||
+ | <p class="wiki_caption">Incubator</p> | ||
</div> | </div> | ||
</div> | </div> |
Latest revision as of 20:06, 20 June 2014
Transformation
a. Take out 50㎕of component cell* kept in 80 degrees below zero and melt it in ice. (We used the compotent cell of One Shot Top 10 Chemically compotent E. coli from Invitrogen.)
b. Add 1㎕ of plasmid DNA** to each of the tube.
c. Mix well and place them in ice for 30 minutes.
d. Place plasmid DNA in the compotent cell and keep it in 42℃ for 30 seconds and place it in ice for 2 minutes.
e. Add 250㎕ of liquid LB broth in each of the tube and grow it in 37℃ shaking incubator for an hour.
f. Leave 100㎕ and dispose.
g. Resuspend the cell, and spread 25㎕ in each of the ampicillin, kanamycin, gentamycin, tetracycline LB solid plate.
h. Spread 25㎕ of competent E. coli in LB solid plate with the same antibiotics.
i. Wait until the E. coli bacteria smears into the plate.
j. Grow it in the 37℃ incubator for a day and observe.
k. Go through miniprep again.
*We used the competent cell from ‘One Shot® TOP10 Chemically competent E. coli’ from Introgen.
**We used the plasmid DNA we got from miniprep previously.