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Minnesota/24 June 2008
From 2008.igem.org
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|3. '''Sequencing primers''' ordered on 6-20-08 were picked up. All primers were diluted to mircomoles according to the following additions of sterile H20: | |3. '''Sequencing primers''' ordered on 6-20-08 were picked up. All primers were diluted to mircomoles according to the following additions of sterile H20: | ||
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|a. All primers were spun down before being opened and H20 added. Primers were then stored at -20C. | |a. All primers were spun down before being opened and H20 added. Primers were then stored at -20C. | ||
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|b. 6 uL reactions containing plasmid to be sequenced and the corresponding primer(s) were set up and submitted for sequencing. | |b. 6 uL reactions containing plasmid to be sequenced and the corresponding primer(s) were set up and submitted for sequencing. | ||
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Revision as of 15:21, 1 July 2008
| 1. Gel Electrophoresis: Used this technique to show plasmid DNA sequence. Materials: |
| a. 50 uL of 1% agarose gel |
| b. Buffer TAE |
| c. One gram of 1% agarose per 100 uL of TAE |
| d. Ethidium bromide (intercalating agent) |
| Problem encountered: electrophoretic gels with 1% agarose had deficient wells |
| Solution: add 0.5 grams more of agarose to the 100uL of TAE buffer |
| 2. Plating from 6-23-08 transformations again. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| a. Since plating of the 6-23 transformations provided no colonies for parts 15-18, the remaining cells from those transformations were replated. 75 uL of cell culture was spread on each of two plates for each culture; plates contained LB media and the corresponding antibiotic. A metal spreading tool was used to spread the culture suspension on the plates, and this was sterilized between each sample by dipping it in 100% ethanol (EtOH) and flaming it. 75 uL cell culture was pipetted on, and spread around plate. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| b. Plates were placed at 37C in an incubator and allowed to grow overnight. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3. Sequencing primers ordered on 6-20-08 were picked up. All primers were diluted to mircomoles according to the following additions of sterile H20:
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