standard Hyperactive TALE-piggyBac Transposase Fusion

Transposon

Transposase interacting with the terminal inverted repeats of a transposon.

piggyBac is a valuable transposon platform upon which to build genetic technologies and now Ye et al. (2015) report creating a modified form of piggyBac transposase with enhanced activity in Bombyx mori.

piggyBac is a reliable transposon with good performance characteristics including good activity and ‘transgene carrying capacity’ but there is room for improvement.  Most notably an increase in transposase activity resulting in a higher frequency of transgenics being produced per germline.

Ye et al. (2015) have fused piggyBac transposase to a TALE (transcription activator-like effector) protein designed to bind to a specific region of the fibroin light chain gene.  Ye et al. make a number of interesting observations.  First, they performed a meta analysis of previous piggyBac integration data in Bombyx mori and calculated the mean integration frequency to be 8.8%.  Second, they create a number of piggyBac transposase constructs with and without a fused TALE protein and test them in two strains of B. mori in conjunction with two vectors.

File:Lifecycle of the Piggybac Transposon System.pngThe presence of TALE consistently led to higher frequencies of piggyBac integration ranging from 1.5-5.7 times higher than controls.  Ye et al. injected TALE-piggyBac transposase mRNA and saw high mortality of embryos injected at 400ng/ul.  They used 200ug/ul in all of their experiments because although it resulted in moderate embryo mortality it also resulted in the most efficient production of transgenics.  The highest frequency of transformation they observed was 63.8%.

Not only did Ye et al. observed higher rates of integration but they also observed a higher number of transgenic individuals coming from germ-lines that produced transgenics.

A TALE linked to a transcription activation domain (VP64) to function as a transcription factor.

A TALE linked to a transcription activation domain (VP64) to function as a transcription factor. Ye et al. have linked the TALE to the piggyBac transposase.

While the TALE-PB fusion seemed to be more active, the reason for this activity is not clear.  None of the recovered piggyBac integration events were in  fibroin light chain where the TALE was programmed to bind, an unexpected result since it was thought that the TALE would tether the PB transposase to the target site and bias integrations to this region of the genome.  It appears that perhaps the increased activity may not be due to tethering of the transposase to the chromosome.  Why the TALE-PB fusion protein is more active than the transpose alone remains unknown but this should not stop anyone from considering using this form of the transposase.

If you are a regular user of piggyBac or are considering it, this paper is worth a look.

Ye, L. et al. TAL effectors mediate high-efficiency transposition of the piggyBac transposon in silkworm Bombyx mori L. Sci. Rep. 5, 17172; doi: 10.1038/srep17172 (2015).

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