Newman Morgan, Lardelli Michael
Discipline of Genetics, The School of Molecular and Biomedical Science, The University of Adelaide, Adelaide, SA, 5005, Australia.
BMC Res Notes. 2010 Nov 4;3:282. doi: 10.1186/1756-0500-3-282.
Transposons are useful molecular tools for transgenesis. The 'sleeping beauty' transposon is a synthetic member of the Tc1/mariner transposon family. Davidson et al. (2003) previously described a vector for zebrafish transgenesis consisting of the inverted repeats of 'sleeping beauty' flanking the gene to be transposed. Subsequently, there have been attempts to enhance the transpositional activity of 'sleeping beauty' by increasing the activity of its transposase. Recently, Mates et al. (2009) generated a hyperactive transposase giving a 100-fold increased transposition rate in mouse embryos.
The aim of this experiment was to determine whether this novel hyperactive transposase enhances transgenesis in zebrafish embryos. Using our previously characterised mitfa-amyloidβ-GFP transgene, we observed an eight-fold enhancement in transient transgenesis following detection of transgene expression in melanophores by whole mount in-situ hybridisation. However, high rates of defective embryogenesis were also observed.
The novel hyperactive 'sleeping beauty' transposase enhances the rate of transgenesis in zebrafish embryos.
转座子是转基因有用的分子工具。“睡美人”转座子是Tc1/水手转座子家族的一个合成成员。戴维森等人（2003年）之前描述了一种用于斑马鱼转基因的载体，该载体由待转座基因两侧的“睡美人”反向重复序列组成。随后，有人尝试通过提高其转座酶的活性来增强“睡美人”的转座活性。最近，梅茨等人（2009年）生成了一种超活性转座酶，在小鼠胚胎中转座率提高了100倍。
本实验的目的是确定这种新型超活性转座酶是否能增强斑马鱼胚胎中的转基因作用。使用我们之前鉴定的mitfa-淀粉样蛋白β-绿色荧光蛋白转基因，通过整体原位杂交在黑素细胞中检测到转基因表达后，我们观察到瞬时转基因作用提高了8倍。然而，也观察到了高比例的胚胎发育缺陷。
新型超活性“睡美人”转座酶提高了斑马鱼胚胎中的转基因率。
