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    • 16. 发明申请
    • METHODS FOR ASSESSING BREAKDOWN PRODUCTS AND STABILITY OF SIRNA AND OTHER TARGET OLIGONUCLEOTIDES
    • 评估SIRNA和其他目标寡核苷酸的突破性产品和稳定性的方法
    • US20090233303A1
    • 2009-09-17
    • US12403597
    • 2009-03-13
    • John F. Thompson
    • John F. Thompson
    • C12Q1/68
    • C12Q1/6869C12Q2525/313C12Q2525/207C12Q2521/501
    • The disclosure provides methods and compositions for assessing by sequencing the type and quantity of in vivo or in vitro breakdown products and stability of nucleic acids such as small interfering RNA (siRNA) and other target oligonucleotides. In general, the disclosed methods involve the use of tester oligonucleotides that comprise a double-stranded region, an optional loop (in the case of hairpin testers), and a single-stranded 3′ overhang that is complementary to the full-length and a shortened target oligonucleotide. A tester is designed so that the 3′ end of a respective target oligonucleotide anneals to the overhang immediately adjacent to the 5′ end of the tester. The juxtaposed ends of the tester and target at adjacent positions allow for a ligase to ligate the chain if there is a match between a tester and its respective target. By sequencing the ligated product in the region of the ligation site, one may determine the sequence of the 3′ end of the target oligonucleotide or of the entire target and its relative amount in the sample.
    • 本公开提供了通过测序体内或体外分解产物的类型和数量以及核酸例如小干扰RNA(siRNA)和其它靶寡核苷酸的稳定性进行评估的方法和组合物。 通常,所公开的方法包括使用包含双链区域,任选环(在发夹测试仪的情况下)的测试者寡核苷酸和与全长互补的单链3'突出端 缩短目标寡核苷酸。 设计一种测试仪,使得各个靶寡核苷酸的3'末端与紧邻测试仪5'端的突出部分退火。 如果测试仪和其各自的目标之间存在匹配,则测试仪和目标位置在相邻位置处并置的端部允许连接酶连接链条。 通过对连接位点的连接产物进行测序,可以确定靶寡核苷酸的3'末端或整个靶标的序列及其在样品中的相对量。