User:Perry/Spring 2007

2/12/07

dissolved 500mg ampicillin salt into 10ml water to prepare 50mg/ml ampicillin stock; aliquotted 1ml into tubes, left in -20dC.

overnight 1ml LB-amp cultures of JLO159, W, H, S, ~4:30pm.

2/13/07

took overnight cultures out ~7:30pm. diluted 1:10, measured OD600.

159- .335, W- .245, H- .231, S- .279

made liquid cultures of 5ml LB-amp + 250ul overnight cultures. incubated 3h, added 5mM IPTG (53ul 0.5M), incubated 2h, diluted 1:10, measured OD600.

159- .106, W- .044, H- .023, S- .057

Aliquotted 1.5mL of H into each tube and proportional volumes of the rest. aliquotted volumes into tubes. centrifuged 8000rpm, 3min. aspirated. reconstituted in 10ul streptavidin aptamer, 10ul B/F hybrid, or 10ul B0/F hybrid.

4uM streptavidin aptamer: 48ul PBS + 2ul streptavidin aptamer (100uM)

4uM B/F hybrid: 46ul PBS + 2ul Bside oligo (100uM) + 2ul Iside oligo (100uM)

4uM B0/F hybrid: 46ul PBS + 2ul DispI oligo (100uM0 + 2ul Iside oligo (100uM)

Wrapped tubes in a napkin, rotated on rotisserie at room temperature for 1h. washed in PBS 3X, reconstituted in 50ul PBS, loaded into clear 96-well plate.