Title:
ONE-POT SINGLE-STRANDED DNA CYCLIZATION AND AMPLIFICATION AND CRISPR/CAS-MEDIATED NUCLEIC ACID MOLECULE DETECTION METHOD
Document Type and Number:
WIPO Patent Application WO/2023/246032
Kind Code:
A1
Abstract:
Disclosed is a one-pot single-stranded DNA cyclization and amplification and CRISPR/Cas-mediated nucleic acid molecule detection method. The method comprises: single-stranded DNA probe cyclization, rolling circle amplification, double-probe CRISPR/Cas shear identification of single-stranded DNA, and single-tube one-pot detection reaction. According to the present invention, in order to mainly solve the problem that cyclization, amplification and shear identification cannot be synchronously carried out, an OPERATOR technology for simultaneous cohesion of cyclization, amplification and shear identification three-step reaction is adopted, and nucleic acid molecules are accurately, sensitively and rapidly detected in an in-vitro single reaction tube system. Compared with a traditional nucleic acid molecule detection method, the present invention has the following advantages that: the method is suitable for sensitive and rapid detection of single-stranded DNA, double-stranded DNA and RNA molecules. Particularly, when detected, RNA is detected directly by one-pot reaction without reverse transcription, and step-by-step reaction is not needed.
Inventors:
YANG LITAO (CN)
ZHU ZAOBING (CN)
GUO YONGKUN (CN)
ZHANG DABING (CN)
ZHU ZAOBING (CN)
GUO YONGKUN (CN)
ZHANG DABING (CN)
Application Number:
PCT/CN2022/140596
Publication Date:
December 28, 2023
Filing Date:
December 21, 2022
Export Citation:
Assignee:
UNIV SHANGHAI JIAOTONG (CN)
International Classes:
C12Q1/6844; C12Q1/6876
Foreign References:
| CN114958978A | 2022-08-30 | |||
| CN113122658A | 2021-07-16 | |||
| CN113943780A | 2022-01-18 | |||
| CN114438262A | 2022-05-06 | |||
| CN114410790A | 2022-04-29 | |||
| CN113308518A | 2021-08-27 |
Other References:
YAN HE, WEN YUNJIE, HAN SONG, HUGHES STEVEN J., ZENG YONG: "One-Pot Endonucleolytically Exponentiated Rolling Circle Amplification by CRISPR-Cas12a Affords Sensitive, Expedited Isothermal Detection of MicroRNAs", BIORXIV, 1 May 2022 (2022-05-01), XP093120004, [retrieved on 20240116], DOI: 10.1101/2022.05.01.490215
王睿璇 (WANG, RUIXUAN): "基于RCA和CRISPR/Cas9的胞外囊泡内miRNA多重检测技术研究 (Non-official translation: Multiplexed Detection of miRNAs in Extracellular Vesicles Based on RCA and CRISPR/Cas9)", 中国优秀硕士学位论文全文数据库 (CHINA MASTER’S THESES FULL-TEXT DATABASE), 15 January 2021 (2021-01-15)
王睿璇 (WANG, RUIXUAN): "基于RCA和CRISPR/Cas9的胞外囊泡内miRNA多重检测技术研究 (Non-official translation: Multiplexed Detection of miRNAs in Extracellular Vesicles Based on RCA and CRISPR/Cas9)", 中国优秀硕士学位论文全文数据库 (CHINA MASTER’S THESES FULL-TEXT DATABASE), 15 January 2021 (2021-01-15)
Attorney, Agent or Firm:
DUAN&DUAN LAW FIRM (CN)
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