Genome engineering: Drosophila melanogaster and beyond.
Genome engineering: Drosophila melanogaster and beyond.
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基因组工程：果蝇果蝇及其他地区。
DOI：
10.1002/wdev.214
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发表时间：
2016-03
期刊：
影响因子：
--
通讯作者：
Hoffman KL
中科院分区：
文献类型：
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作者：
Venken KJ;Sarrion-Perdigones A;Vandeventer PJ;Abel NS;Christiansen AE;Hoffman KL
研究生物现象的一个核心挑战是开发能够通过生殖系传播的核苷酸精确修饰基因组DNA的技术。近年来，这些技术带来了布恩，现在统称为基因组工程。在核苷酸水平上定义的基因组操作使各种逆向工程范式成为可能，为询问不同的生物功能提供了新的机会。这些遗传修饰包括控制去除、插入和替换小的和大的遗传片段。小片段多至几个酶（例如，单个或多个基因座上的单核苷酸突变、小缺失或基因标记）到高达兆碱基分辨率的大片段可以在单个基因座上进行操作，以产生整个染色体臂大部分区域的缺失、重复、倒位或易位。通过同线替换，推测在不同生物体之间功能上正交的染色体部分的专门取代可以提供调节序列之间进化保守性的证据。含有内源性或合成DNA的大型转基因可以整合在确定的基因组位置，允许进化保守的替代证据，复杂的转基因可以用来询问生物现象。精密工程还可用于操纵细胞器的基因组（例如，线粒体）。新型基因组工程范式通常在现有的、易于遗传处理的模式生物中得到加速，主要是因为这些范式可以整合到严格的现有技术基础中。果蝇模型是这些类型研究的理想模型。由于其基因组大小小，只有四条染色体，大量的尖端遗传技术，以及其生命周期短和廉价的维护要求，苍蝇非常适合使用先进的基因组工程进行复杂的遗传分析。因此，在果蝇模型中开发的高度复杂的方法几乎可以用于任何测序的生物体。在这里，我们总结了不同的方法来执行精确的遗传基因组工程使用整合酶，重组酶，和DNA核酸酶在D。黑腹菌
A central challenge to investigating biological phenomena is the development of techniques to modify genomic DNA with nucleotide precision that can be transmitted through the germ line. Recent years have brought a boon in these technologies, now collectively known as genome engineering. Defined genomic manipulations at the nucleotide level enable a variety of reverse engineering paradigms, providing new opportunities to interrogate diverse biological functions. These genetic modifications include controlled removal, insertion, and substitution of genetic fragments, both small and large. Small fragments up to a few kilobases (e.g., single nucleotide mutations, small deletions, or gene tagging at single or multiple gene loci) to large fragments up to megabase resolution can be manipulated at single loci to create deletions, duplications, inversions, or translocations of substantial sections of whole chromosome arms. A specialized substitution of chromosomal portions that presumably are functionally orthologous between different organisms through syntenic replacement, can provide proof of evolutionary conservation between regulatory sequences. Large transgenes containing endogenous or synthetic DNA can be integrated at defined genomic locations, permitting an alternative proof of evolutionary conservation, and sophisticated transgenes can be used to interrogate biological phenomena. Precision engineering can additionally be used to manipulate the genomes of organelles (e.g., mitochondria). Novel genome engineering paradigms are often accelerated in existing, easily genetically tractable model organisms, primarily because these paradigms can be integrated in a rigorous, existing technology foundation. The Drosophila melanogaster fly model is ideal for these types of studies. Due to its small genome size, having just four chromosomes, the vast amount of cutting-edge genetic technologies, and its short life-cycle and inexpensive maintenance requirements, the fly is exceptionally amenable to complex genetic analysis using advanced genome engineering. Thus, highly sophisticated methods developed in the fly model can be used in nearly any sequenced organism. Here, we summarize different ways to perform precise inheritable genome engineering using integrases, recombinases, and DNA nucleases in the D. melanogaster.