即将举行的网络研讨会:实现无需离心的流式细胞术样品制备工作流程的标准化
单细胞测序技术在基础科学研究和临床研究中得到了广 泛应用。单细胞在许多领域发挥着重要作用,在早期诊断、跟踪和个体化治疗中发挥着重要作用。
单细胞测序先不仅仅是对一个细胞进行测序,而是该技术可以对单个细胞的基因组或转录组进行测序,可以理解为单细胞水平的测序。在介绍基本原理之前,让我们试着回答这个问题:为什么要进行单细胞测序?换句话说,单细胞测序能解决哪些传统方法无法解决的问题?世界上没有两片完全相同的树叶。对于多细胞生物来说,细胞之间存在差异。在很多情况下,这只是基因组和转录组上比较轻微的错误,但功能上的差异却有千里之遥。
那么我们如何研究这种遗传信息的异质性呢?传统的测序方法是在多细胞层面进行的,这使得它们以“吃一大锅饭”的形式丢失了异质信息。单细胞测序是这一问题的解决方案。举个更形象的例子:虽然这三个样本之间存在很大的差异,但是通过检测,我们得出了该基因在不同组织中的表达基本相同的结论。
上面显示的异质性被完全忽略了,传统测序方法提供的信息也是多细胞水平的平均信息,而单细胞水平的测序可以充分反映同一细胞群中不同细胞的基因组和转录组状态。单细胞测序技术的出现解决了从混合样本中筛选异质信息的难题,该技术的成熟应用将使生命科学研究向前迈进一大步。那么单细胞测序是如何工作的呢?以单细胞RNA-SEQ为例,简要介绍该技术的原理:分离单细胞,分别构建测序文库,进行测序。这种方法的吞吐量非常低,成本也很高。如上所述,测量几十个细胞需要花费很多钱,而这几十个细胞往往不足以反映真正的科学问题。所以我们关注第二种选择。
基于条形码的单细胞识别。这个想法是在每个细胞的被逆转录时,在测序之前添加一个独特的标记序列。这样,即使结合测序,携带相同标签序列(条形码)的RNA也可以被认为来自同一细胞。通过这种策略,我们可以通过构建数据库一次测量数万个单细胞的信息。
在传统的研究方法中,我们经常根据标记基因和细胞形态来区分不同的细胞类型,这种方法无疑是有争议的。单细胞测序使细胞更加准确和无偏。特别是免疫学、遗传学的研究将产生巨大的影响。分析稀有细胞,特别是那些在特定的空间和时间环境中的细胞。比如从环境中取样的微生物。体外受精胚胎植入前的临床筛选。基于循环细胞的诊断,大力推广循环肿瘤细胞检测新技术。
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