-Targeting-

-Methods- Added 10μl distilled water to wells of J23100 and P0440;distribution kit of iGEM. Then pipetted a couple of times, left it for 5min. After, all collected. NovaBlue T1R used as copetentcell, not ECOS-competent cell.

-Protocol- 1. Melt NovaBlue T1R on the ice. 2. Add 2μl plasumid to NovaBlue T1R, and incubate on the ice for 30 min. 3. Heatshock at 42℃, 30 sec. 4. Incubate on the ice for 2 min. 5. Spread 20-250μl on the plate. 6. Incubate at 37℃ for 12-14 hours.

-Results- E.coli transformed for J23100 and P0440 have been incubating.

-Next experiment- Check the growth of E.coli transformed for J23100 and P0440.(Shimada 1000-) Make LB medium for preculture.(Shimada & Ichikawa 1500-) Preculture E.coli transformed for J23100 and P0440.(Shimada & Ichikawa 2100-)

• Plasmid solution of J23100 and P0440 are keeping in microfuge tubes and in freezer in Cell-biochemistory lab.

Seven plates of LB medium (Amp 50μg/ml)(didn't use) are keeping in cooling room in Biological macinery room.