Optimization of the piggyBac transposon using mRNA and insulators: toward a more reliable gene delivery system.
Optimization of the piggyBac transposon using mRNA and insulators: toward a more reliable gene delivery system.
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DOI：
10.1371/journal.pone.0082559
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Rouleux-Bonnin F
中科院分区：
文献类型：
--
作者：
Bire S;Ley D;Casteret S;Mermod N;Bigot Y;Rouleux-Bonnin F
将转基因整合并稳定表达到细胞基因组中仍然是基于基因的治疗和生物生产目的的主要挑战。虽然转座子载体介导有效的转基因整合，但表达可能受到表观遗传沉默的限制，并且持续的转座酶表达可能介导多个转座循环。在这里，我们评估了piggyBac转座酶信使RNA与遗传隔离转座子的组合的递送，以将转基因与邻近的调控元件分离并稳定表达。从表达水平、转座效率、转基因表达和遗传毒性效应方面对信使RNA和DNA载体的piggyBac转座酶表达进行比较，以校准和确保基于转座的递送系统。信使RNA将转座酶的持久性降低到一个狭窄的窗口，从而减少了副作用，如多余的基因组DNA切割。发现CTF/NF 1和D4 Z4绝缘子两者都从一些转座事件介导更有效的表达。我们的结论是，使用工程piggyBac转座酶mRNA和绝缘转座子提供了有前途的方法，提高质量的整合过程和维持转座子载体的表达。
Integrating and expressing stably a transgene into the cellular genome remain major challenges for gene-based therapies and for bioproduction purposes. While transposon vectors mediate efficient transgene integration, expression may be limited by epigenetic silencing, and persistent transposase expression may mediate multiple transposition cycles. Here, we evaluated the delivery of the piggyBac transposase messenger RNA combined with genetically insulated transposons to isolate the transgene from neighboring regulatory elements and stabilize expression. A comparison of piggyBac transposase expression from messenger RNA and DNA vectors was carried out in terms of expression levels, transposition efficiency, transgene expression and genotoxic effects, in order to calibrate and secure the transposition-based delivery system. Messenger RNA reduced the persistence of the transposase to a narrow window, thus decreasing side effects such as superfluous genomic DNA cleavage. Both the CTF/NF1 and the D4Z4 insulators were found to mediate more efficient expression from a few transposition events. We conclude that the use of engineered piggyBac transposase mRNA and insulated transposons offer promising ways of improving the quality of the integration process and sustaining the expression of transposon vectors.
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影响因子：
5.1
作者：
Gaussin, A.;Modlich, U.;Mermod, N.
通讯作者：
Mermod, N.
DOI：
10.1038/mt.2009.302
发表时间：
2010-04
期刊：
Molecular therapy : the journal of the American Society of Gene Therapy
影响因子：
--
作者：
通讯作者：
--
影响因子：
64.5
作者：
Ding, S;Wu, XH;Xu, T
通讯作者：
Xu, T
影响因子：
23.9
作者：
Lacoste, Arnaud;Berenshteyn, Frada;Brivanlou, Ali H.
通讯作者：
Brivanlou, Ali H.
影响因子：
3.3
作者：
Furushima, Kenryo;Jang, Chuan-Wei;Behringer, Richard R.
通讯作者：
Behringer, Richard R.