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Team:NTNU Trondheim/Journal
From 2011.igem.org
Lab Journal
Thursday 23/6
Lab equipment was prepared: Pipette tips, 1.5 ml tubes, toothpicks, SOC, LA with 100 µg/ml ampicillin/ 100 µg/ml spectinomycin (from 1988). Recipies are given in recipies section.
Friday 24/6
Biobricks were taken out from kit by resuspending them in sterile water followed by transformation to E.coli DH5 alpha. The biobricks that were taken out are given below:
| Abbreviation | Name | Part number | Resistance |
|---|---|---|---|
| P1 | rrnB P1 | BBa_K112118 | Spec |
| lambda pR | lambda pR mod | BBa_R0051 | Amp |
| RFP | TetR + p(TetR)+RFP | BBa_K092600 | Amp |
| Lux | TetR + p(TetR)+RFP+luxI | BBa_K092700 | Amp |
Saturday 25/6
Plates with transformants were moved to fridge.
Sunday 26/6
Transformants were inoculated in 3 ml LB + proper antibiotics to prepare for isolation of plasmid.
Monday 27/6
Plasmids that contained P1, lambda pR, RFP and Lux biobricks were isolated by using miniprep kit from Promega.
Consentrations of the biobricks were measured and were as follows:
| Biobrick | Consentration (ng/µl) |
|---|---|
| P1 | 98.4 |
| lamda pR | 43.8 |
| RFP | 141.8 |
| Lux | 62.8 |
Transformants from Lux/RFP constructs were not red as expected. It was suggested that the phenotype could be explained by presence of TetR in the system. Since tetracycline (Tc) is an inhibitor of TetR we tried to grow the transformants in sublethal consentrations of Tc. E. coli with RFP were grown in 3 ml LB with 0, 0.1, 0.3, 0.6, 1.0, 1.5 and 100 µg/ml Tc.
Tuesday 28/6
It's Jon's birthday! Happy day!
lambda pR i backbone:
· Kutte RFP og RFP + luxI construct med EcoRI og XbaI.
· Kutte lambda pR med EcoRI og SpeI
· Ligate them bitches – win
Kuttet RFP, Lux og Lambda Pr promotor ved bruk av iGem sin protokoll. http://partsregistry.org/Help:Protocols/Restriction_Digest
Alternativ design av system som skal gjøre cellene røde ved tilsats av ppGpp:
