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JULY: WEEK 3
July, 11th
Reactions were incubated at 37°C for three hours while two medium-size agarose gels were prepared according to protocols. In the afternoon gel electrophoresis was performed.
As shown, in figure all clones were positive, so we cut and purified the bands of interest. After gel extraction, cut DNA was quantified:
Then ligations were performed in a final volume of 10 μl:
Ligations were incubated ON at 16°C. July, 12thE9, E10, E11, E12, E13, E14, E15, E16, E17, E18, 19, E20, E21, E22, E23, and E8 were transformed in 100 μl of TOP10 competent cells according to protocols. Plates were incubated ON at 37°C. July, 13thAll plates showed a lot of colonies, except for E13, E16 (one colony),while for E14, E15 no colony was observed. Because of RFP, E21 was red, E22 and E23 a little bit less. Two colonies for E9, E10, E11, E12, E17, E18, E19, E20, E21, E22, E23 plates were picked. 500 ml of LB with Ampicillin were prepared. ??????????????appunti nico?????????????????????-->stock July, 14thA glycerol stock for E21-2, E23-2. E11-1 was prepared. Cultures were saturated; plasmid purification was carried out:
For E9-1, E9-2, E10-1, E10-2, E11-1, E11-2, E12-1, E12-2, E17-1, E18-2, E19-1, E19-2, E21-1, E20-2, E21-1, E21-2, E22-1, E22-2, E23-1, E23-2 PCR was performed. ??????????????????????????????????primer? da scrivere??????????????????????????????????????? <partinfo>BBa_B0021</partinfo>were transformed to test the competence?. In order to screen the DNA, digestions for each ligation was performed for screening.
Reactions were incubated at 37°C for three hours while two medium-size agarose gels were prepared according to protocols. In the afternoon gel electrophoresis was performed.
As shown in figure, all clones were positive, so we cut and purified the bands of interest, except for E9-1,E17-2, E18-1, E18-2, E19-1 which didn' t show any DNA band. FINIRE QUESTA GIORNATA After gel extraction, cut DNA was quantified:
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Team:UNIPV-Pavia/Calendar/July/settimana3
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