The top 20 hits for each such repeat were parsed to a file, and the sequence of each hit was used to BlastX against the amino acid sequences of TE-encoded proteins (http://www.repeatmasker.org/RepeatProteinMask.html database [date last accessed 26 Sep 2011]) to verify that the contig contained the expected target transposition-associated protein-encoding sequences.
Targeted transposition can be a powerful technology for safe transgene integration in human therapeutic applications.
We demonstrated targeted transposition into endogenous matrix attachment regions (MARs) and a chromosomally integrated tetracycline response element (TRE) in cultured human cells, using targeting proteins that bind to the transposon DNA.
Targeting transposition may be possible by coupling a specific DNA binding domain to the SB transposase, but it is not known if this strategy will preserve or disrupt activity of the system.
Targeted transposition was then used to move the P-element into roX1.
We have named this new strategy Targeted Gene Conversion (TGC) to reflect the two-step process required: targeted transposition followed by gene conversion.
A recent study showed that TnsE, a Tn7-encoded factor that targets transposition preferentially to replicating conjugative plasmids, interacts with the β sliding clamp (Parks et al. 2009).
In this study we set out to manipulate the selection of SB's target sites for targeted transposition into predetermined chromosomal regions.
Targeted transpositions are designated as roX1 [MS2-6]tandemandem insertion) or roX1 [MS2-6]replacementement of plArB), followed by the transposition number.
Amplicons spanning the MS2 loop insertion site produce products characteristic of both wild-type roX1 (547 bp) and roX1 MS2-6 (869 bp) from targeted transpositions, but almost 99% of white-eyed offspring produced a single amplicon.
The p[ w +mC GM roX1 MS2-6] transgene was moved into roX1 by targeted transposition, using the roX1 mb710 plArB element as the target site.
