Method for constructing a stable and highly expressed human suspension cell line based on targeting cell membrane surface receptors

By constructing and packaging the retargeting CD19 lentivirus expression vector, using CD19 as a retargeting receptor, the problem of low efficiency of VSVG lentivirus infection suspended cell lines was solved, significantly improving the infection intensity and level, and improving the efficiency of building stable and highly expressed human suspended cell lines.

CN118547010BActive Publication Date: 2025-03-25GUANGZHOU DOUBLLE BIOPRODUCT CO LTD
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Patent Information

Application Number
CN202410705188.3
Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
Filing Date
2024-06-03
Publication Date
2025-03-25
Estimated Expiration
2044-06-03

AI Technical Summary

Technical Problem

In the prior art, the efficiency of using VSVG lentivirus to construct stable and highly expressed human suspended cell lines is low, mainly due to the low expression level of VSVG receptor low density lipoprotein receptor (LDLR) in suspended cell lines, resulting in poor infection effect.

Method used

By constructing a retargeting CD19 lentivirus expression vector and using CD19 as a retargeting receptor, the lentivirus's ability to infect suspended cells is enhanced. Specific steps include constructing the retargeting CD19 lentivirus expression vector pLQ153 or pLQ154, performing retargeting CD19 lentivirus packaging, and infecting target cells through lentivirus to improve the intensity and level of infection.

Benefits of technology

It significantly improves the infection intensity and infection level of lentiviruses on suspended cells, and improves the efficiency of building stable and highly expressed human suspended cell lines.

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Abstract

The present invention discloses a method for constructing a stable high-expression human suspension cell line based on targeting cell membrane surface receptors, and relates to the field of biotechnology. The construction method comprises the following steps: (1) constructing a retargeted CD19 lentiviral expression vector pLQ153 (SEQ ID NO.10) or pLQ154 (SEQ ID NO.12); (2) using a four-plasmid system, retargeting envelope expression vector pLQ153 or pLQ154, packaging plasmid 1, packaging plasmid 2 and envelope plasmid to perform retargeted CD19 lentiviral packaging; (3) adding a lentivirus with a corresponding multiplicity of infection (MOI) to infect the target cells, and adding an infection-promoting agent. The construction method of the present invention has a very good effect on the construction of a high-expression human suspension cell line.
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