Week2: May 27 Out of Lab

Friday, 5/27
LynnMembers present: Celine (for the first half hour), Tam (left at 3:30), Hamed, Cathy, DK, Alex, Katariina
Lab Stuff: (research, plasmid design, protocol design, troubleshooting)
Research into the appropriate promoter:
"Transcription of gesABC is delayed compared with the Au-induced expression of golTS or golB, and requires higher GolS intracellular levels, suggesting a hierarchical activation of the gol regulon to cope with contaminated environments."
In the presence of gold ions: There is a sensitivity/specificity trade-off between using pgolB and pges ( see below)
Sensitivity (i.e. Transcription level in presence of gold(III) ions)Higher (36 folds) i.e. easier to set offLower i.e. harder to set off
Specificity (i.e. Response to Cu(II))Lower i.e. more sensitive to Cu(II)Higher (65 folds) i.e. less sensitive to Cu(II)
Time-course of GolS induction of gesABC and golB transcription in the presence of Au(III) (AuHCl4). The maximum amount of transcription response shows 278 Miller units for gesABC and 10090 Miller units for golB - 36-fold difference. There is an indication that pGesABC was less sensitive to pGolB in the presence of Au(III) as pGesABC transcription level was significantly low in the first 5 hours. Transcription reaches the maximum capacity at 5.8 hours for gesABC and 3.6 hours for GolB.
GesABC::LacZ fused gene expresses no response to the addition of CuSO4 (Cu(II)) in the system. GolB::LacZ fused gene expresses a 65-fold response to the addition of CuSO4 (Cu(II)) in the system.
In general, there is a high level of sensitivity to Au ions in pGolB than in pGesABC. However, pGesABC shows a higher specificity to Au ions than Cu ions in comparision to pGolB which can be induced by both Au and Cu ions.
Creation of golS variants in the feature library using the following table to assist in optimizing the new codons:
E. coli Codon Bias.gif
Administrative: (shipment orders, inventory)
Email Updates: (Correspondance with proffessors, shipping companies, etc)
Followed up regarding lockers
Drafted e-mail to Kayla and sent to Umar for review before sending it out
Tasks to Follow up on - leading into week 3
Promoter debacle (specificity/sensitivity) - All
Luciferase/RFP (reaction with cellulose) - Kat
merR family and Nickle sensors generally (as back up) - Wiki team
Outlining Dr. Pardee's updated protocol - Kat
golS structures (4 versions) - Tam
118 - Kat
113 - Alex
113 + 118 - Alex
CupR - Cathy
Tasks leading into week 3 - v2.jpg
Meeting Notes: