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文件名称：2017ac-单细胞mRNA成像
文件大小：3.85MB
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更新时间：2022-04-25 07:43:30
单细胞成像
ABSTRACT: Ultrasensitive and specific in situ imaging of
gene expression is essential for molecular medicine and clinical
theranostics. We develop a novel fluorescence in situ
hybridization (FISH) strategy based on a new branched
hybridization chain reaction (bHCR) for efficient signal
amplification in the FISH assay and a ligase-mediated
discrimination for specific mutation detection. To our
knowledge, this is the first time that HCR has been realized
for mutation detection in the FISH assay. In vitro assay shows
that the ligation-bHCR strategy affords high specificity in
discriminating single-nucleotide variation in mRNA, and it
generates a highly branched polymeric product that confers more efficient amplification or better sensitivity than HCR. Imaging
analysis reveals that ligation-bHCR generates highly bright spot-like signals for localization of individual mRNA molecules, and
spot signals of different colors are highly specific in genotyping point mutation of individual mRNA. Moreover, this strategy is
shown to have the potential for quantitative imaging of the expression of mRNA at the single-cell level. Therefore, this strategy
may provide a new promising paradigm in developing highly sensitive and specific FISH methods for various diagnostic and
research applications.