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首页> 外文期刊>Cell biology international. >Using a nano-flare probe to detect RNA in live donor cells prior to somatic cell nuclear transfer
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Using a nano-flare probe to detect RNA in live donor cells prior to somatic cell nuclear transfer

机译：在进行体细胞核转移之前，使用纳米火炬探针检测活供体细胞中的RNA

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Many transgenes are silenced in mammalian cells (donor cells used for somatic cell nuclear transfer [SCNT]). Silencing correlated with a repressed chromatin structure or suppressed promoter, and it impeded the production of transgenic animals. Gene transcription studies in live cells are challenging because of the drawbacks of reverse-transcription polymerase chain reaction and fluorescence in situ hybridization. Nano-flare probes provide an effective approach to detect RNA in living cells. We used 18S RNA, a housekeeping gene, as a reference gene. This study aimed to establish a platform to detect RNA in single living donor cells using a Nano-flare probe prior to SCNT and to verify the safety and validity of the Nano-flare probe in order to provide a technical foundation for rescuing silenced transgenes in transgenic cloned embryos. We investigated cytotoxic effect of the 18S RNA-Nano-flare probe on porcine fetal fibroblasts, characterized the distribution of the 18S RNA-Nano-flare probe in living cells and investigated the effect of the 18S RNA-Nano-flare probe on the development of cloned embryos after SCNT. The cytotoxic effect of the 18S RNA-Nano-flare probe on porcine fetal fibroblasts was dose-dependent, and 18S RNA was detected using the 18S RNA-Nano-flare probe. In addition, treating donor cells with 500pM 18S RNA-Nano-flare probe did not have adverse effects on the development of SCNT embryos at the pre-implantation stage. In conclusion, we established a preliminary platform to detect RNA in live donor cells using a Nano-flare probe prior to SCNT.
机译：许多转基因在哺乳动物细胞（用于体细胞核移植的供体细胞[SCNT]）中沉默。沉默与染色质结构的抑制或启动子的抑制有关，并阻碍了转基因动物的生产。由于逆转录聚合酶链反应和荧光原位杂交的缺点，在活细胞中进行基因转录研究具有挑战性。纳米耀斑探针提供了一种检测活细胞中RNA的有效方法。我们使用管家基因18S RNA作为参考基因。这项研究旨在建立一个平台，以便在SCNT之前使用Nano-flare探针检测单个活体供体细胞中的RNA，并验证Nano-flare探针的安全性和有效性，从而为挽救转基因中沉默的转基因提供技术基础。克隆的胚胎。我们研究了18S RNA-Nano-flare探针对猪胎儿成纤维细胞的细胞毒性作用，表征了18S RNA-Nano-flare探针在活细胞中的分布，并研究了18S RNA-Nano-flare探针对猪胎儿成纤维细胞的影响。在SCNT之后克隆了胚胎。 18S RNA-Nano-flare探针对猪胎儿成纤维细胞的细胞毒性作用是剂量依赖性的，使用18S RNA-Nano-flare探针可检测18S RNA。此外，在植入前阶段用500pM 18S RNA-Nano-flare探针处理供体细胞对SCNT胚胎的发育没有不利影响。总之，我们建立了一个初步平台，用于在SCNT之前使用Nano-flare探针检测活体供体细胞中的RNA。

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