Three new D–π–A type dyes 1–3 comprising triphenylamine as the core and [12]aneN₃ or benzothiazole (Bz) as branches were designed as two-photon luminescent non-viral gene vectors with tracking ability. Compound 1 with one Bz and one [12]aneN₃ branches as well as a heavy atom, I, has higher toxicity and lower transfection efficiency. Compound 2 containing two [12]aneN₃ branches shows a good mitochondria targeting property and can condense DNA at 5 μM, in particular it exhibits 1.70 times higher red fluorescent gene (RFG) transfection efficiency than Lipofectamine 2000 in the presence of cell penetrating peptides (CPPs). Compound 3 with two Bz-branches and one [12]aneN₃ moiety shows the largest two-photon absorption cross section of 309 GM at 760 nm among the three dyes and is able to localize in mitochondria with an overlap coefficient of 0.99. With the advantages of strong two-photon absorption, good DNA condensation, and mitochondria targeting ability, compound 2 was used to monitor the process of cellular uptake, DNA translocation and release, RFG expression in cells, and RFG expression in a rat tumor using a two-photon microscope. To the best of our knowledge, this study represents the first example of a mitochondria targeting two-photon fluorescent organic molecule in gene transfection in vitro and in vivo.

中文翻译：

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靶向线粒体的两光子荧光分子用于基因转染和生物跟踪†

以三苯胺为核心和[12] aneN ₃或苯并噻唑（Bz）为分支的三种新型D–π–A型染料1–3被设计为具有跟踪能力的双光子发光非病毒基因载体。具有一个Bz和一个[12] aneN ₃分支以及重原子I的化合物1具有更高的毒性和更低的转染效率。含有2个[12] aneN ₃分支的化合物2具有良好的线粒体靶向性能，可以浓缩5μM的DNA，特别是在存在细胞穿透肽的情况下，它的红色荧光基因（RFG）转染效率是Lipofectamine 2000的1.70倍（ CPPs）。化合物3具有两个Bz分支和一个[12] aneN ₃部分的化合物在三种染料中显示最大的309 GM在760 nm的双光子吸收截面，并且能够以0.99的重叠系数定位于线粒体中。化合物2具有较强的双光子吸收，良好的DNA凝聚和线粒体靶向能力的优点，因此使用2号化合物来监测细胞摄取，DNA易位和释放，细胞中RFG表达以及大鼠肿瘤中RFG表达的过程。双光子显微镜。据我们所知，这项研究代表了在体外和体内基因转染中靶向双光子荧光有机分子的线粒体的第一个实例。