From 2007.igem.org

yesterday -- tomorrow

Cloning

All our plates were covered with a lawn of bacteria... We think our LB was contaminated... But we had 5µl of our ligations mix remaining (which stayed on the bench ON), so we decided to try and transform them again! We'll just see what happens!

Analysis of the sequences

We sequenced the few positives clones we got from the pTOPO cloning of the DapA PCR products.

• Lox66-DapAColi: 2 clones, the biobrick prefix is missing in both
• Lox66-DapASubtilis: 2 clones, lox66 is mutated in both, Pst1 is missing in both
• RBS-DapAColi: 3 clones ok
• RBS-DApASubtilis: 3 clones, 2 clones seem ok

The oligos used to amplify lox66-DapA were 81bp... They probably were not very good up to the end. The end of the PCR products are often missing. This might explains why we had so much difficulties in cloning them. But at least, now we got them and we can go on.