Part:BBa_K1189000
TALE-A with a his tag under a lacI promoter
TALA(designed by Slovenia's 2012 iGEM team) was built down stream of BBa_ J04500, an IPTG inducible promoter with RBS. Since this gene was used in eukaryotic cells by Slovenia's iGEM team, it had a Kozak sequence at the start of it. We removed this Kozak sequence to allow for the expression of the protein in E. coli
Figure 6: TALE capture assay was done with TALE B (
BBa_K1189001
)and TALE A B-lac fusion (
BBa_K1189031
) and DNA (
BBa_K1189006
)
with both target sequences. If capture is successful, the B-lac is present in the well giving a colour change from pink to yellow when subjected to benzylpenicillin substrate solution within 20 minutes. The only wells that change colour are the first four wells which contain TALE B, specific DNA, and TALE A B-lac fusion and our positive control wells which are the controls for our fusion TALE A B-lac protein and our positive recombinant b-lac. All our other controls including our test using a non-specific sequence of DNA remained pink . This preliminary characterization data demonstrates that the TALEs are able to bind to DNA with specificity. Additionally it also shows that our system of capturing DNA with two detector TALEs and then subsequent reporting of the DNA’s presence works.
This assay shows that we can capture our target DNA with two detector TALEs with specificity . Additionally, we can report whether that DNA has been captured and is present in the sample, which is a very important concept for our sensor system.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 2654
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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