HyperActive piggyBac Transposase for Transgenesis

Nathaniel Grubbs, Ph.D., is a regular contributor to Technology Topics. He is a postdoctoral research in the Department of Entomology, North Carolina State University

Eckermann et al. (2018) recently published a report in Insect Biochemistry and Molecular Biology demonstrating the efficacy of hyperactive piggyBac transposase for creating transgenic insects, which may be of interest to researchers looking to initiate transgenic projects in ‘non-model’ insect species.

Derived from the cabbage looper moth, piggyBac has become a common transposon in transgenic work, since it has proved active in a variety of species, including vertebrates. However, this hasn’t stopped researchers from trying to increase its efficiency beyond the wild-type activity. For this reason, a hyperactive version of the piggyBac transposase was developed (Yusa et al., 2011). Two versions were created, optimized for mammalian and insect codon biases, which Eckermann refers to as mhyPBase and ihyPBase, respectively. Shortly after its development, ihyPBase was put to the test by Wright et al. (2013) in Drosophila melanogaster and a mosquito, Aedes aegypti. Unfortunately, Wright and her colleagues encountered heightened sterility in both species, which seemed, in part, to be caused directly by the transposase, independent of transformation. Worse still, this sterility was not accompanied by any

increase in transformation efficiency relative to the wild-type protein.

Eckermann and colleagues had a very different experience with ihyPBase, so decided to share their own results, testing both mhyPBase and ihyPBase against the original piggyBac transposase in double-blind transgenic experiments. They also used three different species: D. melanogaster, another dipteran from a different family, Ceratitis capitata, and a beetle species, Tribolium castaneum. In all three species, Eckermann et al. found enhanced transgenesis from both hyperactive transposases relative to the original. Interestingly, they found little evidence that codon bias was important, since the transformation rate of mhyPBase was usually comparable to that of ihyPBase. Importantly, they found no evidence of a decrease in fertility after injections of either hyperactive transposase versus the wild-type protein.

While it may be surprising that two different groups could generate such different results, Eckermann does remind readers that a number of mechanical caveats influence the outcomes of microinjections. Indeed, Eckermann and colleagues confess in their Materials and Methods that they managed their injection volumes to enhance survival rather than transgenesis, and both groups used distinct concentrations of helper and donor plasmids in their experiments. Eckermann’s group even used injection buffer, a potentially influential element that Wright does not discuss using. In the end, while both groups appear to use proper controls, their experiments are sufficiently different to offer a plethora of potential explanations for their distinct outcomes.

Medfly, Cerratitis capitata
By Alvesgaspar – https://commons.wikimedia.org/w/index.php?curid=5027949

Tribolium castaneum

Most important, however, is that Eckermann et al. demonstrate that the hyperactive piggyBac transposase is capable of performing above and beyond the wild-type version. These findings will be of particular use to researchers starting transgenic programs with under-studied insects, particularly those that are difficult to rear and/or microinject. Hyperactive piggyBac transposase could prove helpful in overcoming species-specific limitations to transgenesis.

Eckermann KN, Ahmed HMM, KaramiNejadRanjbar M, Dippel S, Ogaugwu CE, Kitzmann P, Isah MD, Wimmer EA. 2018. Hyperactive piggyBac transposase improves transformation efficiency in diverse insect species. Insect Biochemistry and Molecular Biology 98: 16-24 doi: 10.1016/j.ibmb.2018.04.001.



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