Method and kit for rapidly screening gene editing positive monoclone based on qPCR (quantitative polymerase chain reaction)
By designing specific primers and probes based on qPCR, combined with single-cell lysis and dual-channel detection, the problems of inaccurate screening and insufficient throughput of traditional methods have been solved, enabling rapid and accurate screening of gene-edited positive monoclonal clones to meet clinical needs.
CN121852520APending Publication Date: 2026-04-14HANGZHOU WUWEN QINGXIN ARTIFICIAL INTELLIGENCE BASIC TECHNOLOGY RESEARCH CO LTD
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Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-01-06
- Publication Date
- 2026-04-14
AI Technical Summary
Technical Problem
Traditional methods for screening gene-edited positive monoclonal clones are inaccurate and difficult to meet the needs of large-scale screening. Existing qPCR technology lacks specific detection solutions for gene-editing events.
Method used
Using a qPCR-based approach, specific primers and probes were designed, and combined with single-cell lysis and dual-channel detection, gene-editing-positive monoclonal clones were directly screened. Editing efficiency was determined by Ct value differences, and multiplex detection was performed using a matching kit.
Benefits of technology
It enables rapid and accurate screening of gene-edited positive monoclonal clones, meeting clinical requirements, and significantly improving screening throughput without relying on screening tags.
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Figure CN121852520A_ABST
Abstract
The invention discloses a method and a kit for rapidly screening gene editing positive monoclone based on qPCR (quantitative polymerase chain reaction), and belongs to the field of gene engineering. The method comprises the following steps: preparing a monoclonal sample: preparing a single-cell suspension from cells subjected to gene editing, and carrying out single-cell sorting and culture until monoclone is formed; collecting monoclonal cells, and directly cracking by using a cracking solution to prepare a qPCR template; qPCR detection: carrying out qPCR amplification and detection on the sample by using a specific primer and a probe designed aiming at a target gene editing event; and result judgment: judging whether the monoclone is gene editing positive clone or not by analyzing the Ct value of the qPCR. According to the present invention, the specific primers and the probe of the target gene editing site are designed, and the qPCR is combined to directly detect the genome variation of the monoclonal cell so as to achieve the early rapid screening;
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