IGEM:MIT/2006/Notebook/2007-8-3

=Progress on Media Front=

MG1655 grew up in all three medias after only ~12 hours of growth. The other colonies look clear, but we will give them time to grow.

=Meeting at 2:00=

On what to do about device characterization for phase-sensitive wintergreen.

Catharsis from Meeting

I should...

1. Check sizes for tetR, kanR, protein in inverter to look at for protein gel (make sure they are not ~39 kDa)

2. Check rare codon usage in all proteins (BSMT, pCHA, pCHB, ATF1, THI3, BAT2, C0040's protein)

3. Do a protein gel of GFP constructs (exponential phase and stationary phase for J45995, J45996, R0040.E0840, and control)

4. Add untransformed TOP10 cells as control in media experiments and transfer 700 and 800 to MG1655

5. Put J45181 and J45250 in same cell and see if they smell one way or another in stationary and exponential phase

What I can do Today

1. Just do it- kanR- 27.17 kDa, ampR- 28.67 kDa, tet repressor (inverter component)- 22 kDa, tetR- 39.7 kDa (UNFORTUNATE)

2. Just do it

3. Make LCs of the GFP constructs to grow up to exponential phase and prepare protein samples tomorrow- DONE

4. Grow up MG1655 LC tonight to make competent tomorrow- DONE and grow up TOP10 cells in EZ media- DONE

5. Many things...


 * Obtain 3K3 EP cut from Nishant and Karen- DONE


 * Obtain 3C5 EP cut from Reshma- DONE


 * Cotransform J45181 and J45250- HOLD OFF UNTIL TOMORROW


 * Make LB AMP TET KAN, LB AMP TET CHL, LB CHL, and LB KAN plates- DONE


 * Digest J45250 with E and P and J45600 (to be cotransformed with J45800 1AK3) with E and P (to be cotransformed with J45181 1AT3)- DONE


 * Ligate and transform J45250 3K3 and J45600 3C5- HOLD OFF UNTIL TOMORROW

=Other things to Do=

1. Grow up MG1655 cells to make chemically competent tomorrow. I think that it may be worth moving J45800 and J45700 into MG1655, so we don't have to wait as long to get results from these media experiments- DONE

2. Grow up 25-mL cultures to do extraction efficiency experiments with the GC tomorrow- SCRAP THIS FOR NOW

3. Email Meagan about IK J45200 and J45120 cells again- DONE

4. Reply to Professor Tribe, thanking him for his advice/telling him what we are doing- DONE

5. Book 68-574 for meeting- DONE