Streamlined and quantitative detection of chimerism in mouse tissue using digital PCR
Streamlined and quantitative detection of chimerism in mouse tissue using digital PCR
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使用数字 PCR 简化和定量检测小鼠组织中的嵌合状态
DOI：
10.1101/2020.11.04.368944
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发表时间：
2020
期刊：
影响因子：
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通讯作者：
Nakauchi Hiromitsu
中科院分区：
文献类型：
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作者：
Suchy Fabian P.;Nishimura Toshiya;Wilkinson Adam C.;Higuchi Maimi;Bhadury Joydeep;Nakauchi Hiromitsu
从发育生物学到癌症研究，动物嵌合体被广泛用于生物医学发现。然而，嵌合和镶嵌组织中混合细胞类型的准确定量具有挑战性。在这里，我们已经开发并表征了一种液滴数字聚合酶链式反应单核苷酸识别试验，以检测白化和非白化小鼠品系之间的嵌合体。此外，我们已经证实，这种分析方法与大多数器官的粗裂解物兼容，大大简化了样品准备。与现有方法相比，这种嵌合体检测方法具有许多额外的优点，包括其健壮的性质、最小的技术偏差以及报告准备样本中的细胞总数的能力。重要的是，这里开发和讨论的概念很容易适应于其他基因组座位，以准确地测量任何组织中的混合细胞群体。
Animal chimeras are widely used for biomedical discoveries, from developmental biology to cancer research. However, the accurate quantitation of mixed cell types in chimeric and mosaic tissues has challenges. Here, we have developed and characterized a droplet digital PCR single-nucleotide discrimination assay to detect chimerism among albino and non-albino mouse strains. In addition, we have validated that this assay is compatible with crude lysate from most organs, drastically streamlining sample preparation. This chimerism detection assay has many additional advantages over existing methods including its robust nature, minimal technical bias, and ability to report the total number of cells in a prepared sample. Importantly, the concepts developed and discussed here are readily adapted to other genomic loci to accurately measure mixed cell populations in any tissue.