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    • 2. 发明申请
    • ENHANCED METHODS OF RIBONUCLEIC ACID HYBRIDIZATION
    • 增强抗核酸杂交的方法
    • WO2015085194A1
    • 2015-06-11
    • PCT/US2014/068835
    • 2014-12-05
    • THE BROAD INSTITUTE, INC.THE GENERAL HOSPITAL CORP
    • BHATTACHARYYA, RobyHUNG, DebPATEL, Milesh
    • C12Q1/68
    • C12Q1/6832C12Q2523/113C12Q2527/101C12Q2527/113
    • This disclosure relates to a method for increasing the hybridization efficiency of a probe and a target RNA in a sample, for example to identify a particular RNA present in the sample. The method includes heating a lysate sample comprising at least one target RNA, such as a tRNA, mRNA or rRNA, at a temperature of about 95°C for a time sufficient to interfere with secondary structure of the RNA, wherein the time is short enough, such that the RNA in the cell lysate sample are not significantly degraded, and wherein the lysate comprises a cell lysis buffer comprising a chemical denaturant. To detect a target RNA in the lysate, the lysate is contacted with at least one detectable probe, such as a labeled probe, designed to specifically hybridize to the target RNA in the lysate.
    • 本公开涉及提高样品中探针和靶RNA的杂交效率的方法,例如以鉴定样品中存在的特定RNA。 该方法包括在约95℃的温度下加热包含至少一种靶RNA(例如tRNA,mRNA或rRNA)的裂解物样品一段足以干扰RNA二级结构的时间,其中时间足够短 使得细胞裂解物样品中的RNA没有显着降解,并且其中裂解物包含包含化学变性剂的细胞裂解缓冲液。 为了检测裂解物中的靶RNA,将裂解物与设计为与裂解物中的靶RNA特异性杂交的至少一种可检测的探针(例如标记的探针)接触。