Versatile synthesis and rational design of caged morpholinos.
Versatile synthesis and rational design of caged morpholinos.
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DOI：
10.1021/ja809933h
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发表时间：
2009-09-23
影响因子：
15
通讯作者：
Chen JK
中科院分区：
文献类型：
--
作者：
Ouyang X;Shestopalov IA;Sinha S;Zheng G;Pitt CL;Li WH;Olson AJ;Chen JK
胚胎发生是由以刻板方式动态执行的遗传程序调节的，并且破译这些分子机制需要以类似的空间和时间精度控制胚胎基因功能的能力。化学技术可以使这样的遗传操作，如使用笼状吗啉代（cMO）寡核苷酸的例子，以在斑马鱼和其他光学透明的生物体的时空控制的基因。在这里，我们报告的优化方法的设计和合成的发夹cMO纳入二甲氧基硝基苄基（DMNB）为基础的双功能连接器，允许cMO组装在只有三个步骤，从市售试剂。使用这种简化的程序，我们系统地制备了具有不同结构构型的cMO，并研究了这些试剂的体外热力学性质与其体内活性之间的关系。通过这些研究，我们建立了cMO设计的一般原则，并成功地将其应用于几个发育基因。我们优化的合成和设计方法也使我们能够制备下一代cMO，其包含基于溴羟基喹啉（BHQ）的连接体用于双光子解锁。总的来说，这些进展确立了cMO技术的普遍性，并将促进这些化学探针在体内用于功能基因组研究。
Embryogenesis is regulated by genetic programs that are dynamically executed in a stereotypic manner, and deciphering these molecular mechanisms requires the ability to control embryonic gene function with similar spatial and temporal precision. Chemical technologies can enable such genetic manipulations, as exemplified by the use of caged morpholino (cMO) oligonucleotides to inactivate genes in zebrafish and other optically transparent organisms with spatiotemporal control. Here we report optimized methods for the design and synthesis of hairpin cMOs incorporating a dimethoxynitrobenzyl (DMNB)-based bifunctional linker that permits cMO assembly in only three steps from commercially available reagents. Using this simplified procedure, we have systematically prepared cMOs with differing structural configurations and investigated how the in vitro thermodynamic properties of these reagents correlate with their in vivo activities. Through these studies, we have established general principles for cMO design and successfully applied them to several developmental genes. Our optimized synthetic and design methodologies have also enabled us to prepare a next-generation cMO that contains a bromohydroxyquinoline (BHQ)-based linker for two-photon uncaging. Collectively, these advances establish the generality of cMO technologies and will facilitate the application of these chemical probes in vivo for functional genomic studies.
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影响因子：
2.9
作者：
MARKY, LA;BRESLAUER, KJ
通讯作者：
BRESLAUER, KJ
影响因子：
4.7
作者：
Bonnet, Dominique;Ilien, Brigitte;Hibert, Marcel
通讯作者：
Hibert, Marcel
影响因子：
3.4
作者：
Nguyen, Quan N.;Chavli, Rajesh V.;McMaster, Gary K.
通讯作者：
McMaster, Gary K.
影响因子：
4.8
作者：
Monroe, WT;McQuain, MM;Haselton, FR
通讯作者：
Haselton, FR
影响因子：
48
作者：
Guo, Yan-Ming;Chen, Shiuhwei;Li, Wen-hong
通讯作者：
Li, Wen-hong