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电穿孔法向植物转移基因:方法与应用。

Gene transfer to plants by electroporation: methods and applications.

机构信息

Department of Biology, Faculty of Science and Arts, Marmara University, Goztepe, 34722, Istanbul, Turkey.

Department of Biology, Faculty of Science, Kyrgyz-Turkish Manas University, 720038, Bishkek, Kyrgyzstan.

出版信息

Mol Biol Rep. 2020 Apr;47(4):3195-3210. doi: 10.1007/s11033-020-05343-4. Epub 2020 Apr 2.

DOI:10.1007/s11033-020-05343-4
PMID:32242300
Abstract

Developing gene transfer technologies enables the genetic manipulation of the living organisms more efficiently. The methods used for gene transfer fall into two main categories; natural and artificial transformation. The natural methods include the conjugation, transposition, bacterial transformation as well as phage and retroviral transductions, contain the physical methods whereas the artificial methods can physically alter and transfer genes from one to another organisms' cell using, for instance, biolistic transformation, micro- and macroinjection, and protoplast fusion etc. The artificial gene transformation can also be conducted through chemical methods which include calcium phosphate-mediated, polyethylene glycol-mediated, DEAE-Dextran, and liposome-mediated transfers. Electrical methods are also artificial ways to transfer genes that can be done by electroporation and electrofusion. Comparatively, among all the above-mentioned methods, electroporation is being widely used owing to its high efficiency and broader applicability. Electroporation is an electrical transformation method by which transient electropores are produced in the cell membranes. Based on the applications, process can be either reversible where electropores in membrane are resealable and cells preserve the vitality or irreversible where membrane is not able to reseal, and cell eventually dies. This problem can be minimized by developing numerical models to iteratively optimize the field homogeneity considering the cell size, shape, number, and electrode positions supplemented by real-time measurements. In modern biotechnology, numerical methods have been used in electrotransformation, electroporation-based inactivation, electroextraction, and electroporative biomass drying. Moreover, current applications of electroporation also point to some other uncovered potentials for various exploitations in future.

摘要

发展基因转移技术使生物体的基因操作更加高效。基因转移的方法主要分为两类:自然转化和人工转化。自然转化方法包括接合、转座、细菌转化以及噬菌体和逆转录病毒转导,包含物理方法;而人工转化方法可以通过物理手段改变和将基因从一个生物体的细胞转移到另一个生物体的细胞中,例如,利用弹道式转化、微注射和宏注射、原生质体融合等方法。人工基因转化也可以通过化学方法进行,包括磷酸钙介导、聚乙二醇介导、DEAE-葡聚糖介导和脂质体介导的转移。电方法也是一种通过电穿孔和电融合来转移基因的人工方法。相比之下,在所有上述方法中,由于其高效率和更广泛的适用性,电穿孔被广泛应用。电穿孔是一种通过在细胞膜中产生瞬时电孔来实现的电转化方法。根据应用的不同,该过程可以是可逆的,其中膜中的电孔可以重新封闭,细胞保持活力,也可以是不可逆的,其中膜无法重新封闭,细胞最终死亡。通过开发数值模型来迭代优化场的均匀性,可以解决这个问题,同时考虑细胞的大小、形状、数量和电极位置,并辅以实时测量。在现代生物技术中,数值方法已被用于电转化、基于电穿孔的失活、电萃取和电穿孔生物质干燥。此外,电穿孔的当前应用也指出了未来在各种开发中的一些未被发现的潜力。

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