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Team:Illinois/MicA
From 2009.igem.org
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MicA Target-GFP Fusion
Primers Used: for the sRNA gene we used Forward Primer: GAA AGA CGC GCA TTT GTT ATC
Temp: (4*9) + (2*12) = 60 degrees
Second homology sequence: TTC CAG CCA CAC CGC AAA CGG ((TTCGGTATCA))
Reverse complement: CCG TTT GCG GTG TGG CTG G Temp: (4*13) + (2*6) = 64 degrees Cut site and overhang: GTTTTT TCTAGA
Reverse Primer: GTTTTT TCTAGA CCG TTT GCG GTG TGG CTG G
For the Target Sequence we used the primers from the Urban and Vogel paper: JV-0432 and JV-0433
June 16
We used PCR to extract the sRNA gene: MicA and target sequence : ompA. from the e.coli chromosome. We then ran a gel to make sure that we had the right DNA fragments. Our results corresponded to our predictions.
June 17
We are completing a digestion of MicA, OmpA, and OmpF. Following the digestion we will incubate with SAP and then run a gel to verify the digestion. We will then extract the DNA for the sRNA target sequence and sRNA gene.
