Lab Sep 18 2009

Well, no flourescence from K235013 or 14, and 14 should definitely have worked. We also tested K235009 as a negative control and E0240 and saw nothing.

We first used Dr. Pearson's microscope and found cells but no flourescence.

We then tried Dr. Burke's microscope and found tiny artifact flourescence on K9 and K14, our negative control and positive respectively, thus meaningless. We didn't bother with K13 and E0240 on that microscope.

2:00pm Kyliah

The way I see it, we never attempted to sequence anything, so that might be the problem. Maybe we should see about PCR'ing up our inserts somehow and ensuring they're the right things.

The BioBrick primers VF2 and VR should work with all three of the plasmids from the P1010s we've been using - the chloramphenicol and kanamycin ones have one single base mismatch, and it's a T where a C was expected, so the primer should still work just fine. They're both 20nt long, so it should be simple to order some... A third to possibly order is the last 20nt of the ribolock we're using, so that we can properly sequence something that's normally self-complementary. If this properties calculator is accurate, the last 20nt should have a similar Tm to the two primers.

5:00pm

Looking at the registry, part of our fails might have been because key3d is screwing up all over the Registry sequencing. Part J23070 was the second-best key for Berkeley even without a terminator. If we rehydrate this part (plate 1, well 16O, pSB1A2-derived plasmid with pTet- -RFP) and then construct a double-terminator onto the end (B0015) we might have a part that's even more effective than the key we have now was claimed to be.

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