May 25
☀ Yi Han ☀
❱❱❱ Received 2 bacterial stab cultures, EsaR/I plasmid from addgene (CHL) and BBa_K299812 from iGEM HQ.
❱❱❱ Streaked out on plates with amp.
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May 26
☀ Adrian ☀
❱❱❱ Transformed:
+Kit plate 1 9N Ba_K763002 chl
+Kit plate 4 13L BBa_E0040 amp
❱❱❱ DNa was received in powder form in plates, and resuspended in 10ul ultrapure H2O respectively. Plates were stored in -20/
☀ Yi Han ☀
❱❱❱ Plates cracked in incubator as they dried from lack of humidity.
❱❱❱ Transfer to small incubator with beaker of water for humidity no single colonies for inv plasmid -> streak again.
❱❱❱ Wrong antibiotic for EsaR/I plasmid → streak out again
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May 28
☀ Yi Han ☀
❱❱❱ Inoculate single colony of invasin plasmid carrying bacteria in 3mL LB+amp
❱❱❱ Transformation of 13L repeated with 1ul of DNA.
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May 29
☀ Xin Yi ☀
❱❱❱ No colonies grew for 13L on all plates → adjust incubator, make new media
❱❱❱ Miniprep of inoculated bacteria for invasin plasmid, incubate sample in 37degC for 1h 20min:
| RE of invasin plasmid |
RE control |
| Rsal 1ul |
Rsal 0 ul |
| EcoRI 1 ul |
EcoRI 0 ul |
| INv plasmid 7.5ul |
Inv plasmid 7.5 ul |
| H2O 35.5 ul |
H2O 37.5 ul |
| NEB buffer 4.5 ul |
Buffer 4.5 ul |
❱❱❱ Repeat transformation of 13L with 1ul
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May 30
☀ Xin Yi & Yi Han ☀
❱❱❱ Miniprep of EsaR/I plasmid for 4 colonies
❱❱❱ Restriction digest for EsaR with XbaI/BamHI:
RE reaction
5ul plasmid
5ul buffer
1ul XbaI
1ul BamHI
Add H2O to 50ul
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