ClingE coli Lets get together Quorumsensing Stephanie Lo

Cling-E. coli Let’s get together: Quorum-sensing Stephanie Lo, Perry Tsai July 9, 2007

Random peptide display and screening Lpp. Omp. A Transformation + MACS (target with magnetic bead) Expression FACS (target with fluorescent tag)

Bringing bacteria together Target (eg Nickel)yo ‘sup lux. R lux. I OHHL

Test constructs OHHL lux. R BBa_I 13263 BBa_T 9002

Growth data for log phase determination

Fluorescence response to OHHL induction GFP YFP 100 n. M OHHL 10 n. M OHHL Non-induced No lux. R

Bio. Bricks, assemble! lux. R lux. I Lux. I generator Lux. R generator / Lux. I receiver / Reporter lux. I/lux. R Lux. I/Lux. R generator and receiver

Looking into new Bio. Bricks Lux. I amplifier Lux. R generators with different promoters Lux. I amplifier and reporter Lux. I receiver with different reporter

In other Lpp. Omp. A news… Lpp-Omp. A PDZ Western blot PDZ domain (+) GFP-Mek LO-PDZ IPTG(-) LO-PDZ IPTG(+) 0 1 2 1+2

Synthesized Lpp. Omp. A (s. LO) Original Lpp. Omp. A Bio. Brick-assembled Lpp. Omp. A ACTAGA Thr. Arg Mixed Synthesized Lpp. Omp. A ACTAGA Thr. Arg Mixed

This week’s agenda l l l FACS appointment on July 11 Checking out ligations and new Bio. Bricks More fluorescence assays, possibly with new constructs Construction of GFPmek for PDZ binding Preparing s. Lpp. Omp. A for future assembly