Illinois/30 July 2008

LAB WORK

Antibody/GPCR Fusion
 * Autoclaved Glass Beads and Eppendorf tubes

Receptor Tyrosine Kinase method

Assembly PCR: amplify antibody gene fragments and then assemble them, light and heavy chains

Master mix already contains MgCl2; should have added extra to some tubes.

PCR program: 1. 94 degrees 5 min 2. 94 degrees 1 min 3. 50 degrees 1 min 4. 72 degrees 1 min 5. GOTO 2. 39 cycles 6. HOLD at 4 degrees

1.5% agarose gel made, 0.75g agarose in 50ml 0.5X TBE w/10ul EtBr.

20ul of PCR produts loaded on gel w/4ul 6X loading buffer, run at 200V for about half an hour.

All lanes had lots of DNA at ~375bp, PCR worked.