Modular Genetically Engineered Cells for Targeted Transgene Integration
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Conventional methods for integrating multiple transgenes in cells are inefficient, lack target site control, and have limited transgene sequence length, making them unsuitable for rapid experimentation and genetic modification.
Innovation Solution
A method for creating a cell line with multiple landing sites at safe harbor locations, allowing for the integration of multiple transgenes in a single shot using site-specific recombinase (SSR) and prime editing technologies, enabling efficient and targeted integration of genes of interest.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods are used for integrating multiple transgenes, then the process can be performed with existing technology, but the efficiency is low and multiple cell selections are required
Solution Approach 1:
The patent pre-integrates landing sites with SSR attachment sequences into the host cell genome at safe harbor locations before transgene integration. This preliminary action enables subsequent rapid integration of multiple transgenes via SSR recombination without requiring multiple rounds of cell selection, as the landing sites are already in place to receive the transgenes.
Solution Approach 2:
The patent introduces landing sites as intermediary elements between the host genome and transgenes. These landing sites contain SSR attachment sequences that serve as mediators for efficient transgene integration through site-specific recombination, eliminating the need for direct random integration and multiple selection processes.
2Manufacturing precision
If conventional transgene integration methods are used, then existing technology can be applied, but target site control is lacking
Solution Approach 1:
The patent integrates landing sites at specific safe harbor locations in the host genome (such as AAVS1, CCR5, or Rosa26 loci) rather than allowing random integration. Each landing site is precisely positioned at a predetermined genomic location with appropriate SSR attachment sequences, enabling controlled and targeted transgene integration while maintaining genomic stability.
Solution Approach 2:
The patent divides the integration system into modular components: landing sites with SSR attachment sequences integrated at specific genomic locations, and transgenes flanked by complementary SSR sequences. This segmentation allows independent design and optimization of each component while maintaining overall system control and precision.
3Length of stationary object
If conventional methods are used for transgene integration, then standard procedures can be followed, but transgene sequence length is limited
Solution Approach 1:
The landing sites act as intermediaries that can accommodate transgenes of varying lengths. The SSR recombination mechanism facilitates integration of large transgene cassettes without the size constraints of viral vectors or other delivery systems, as the recombination process itself is not limited by transgene length.
Solution Approach 2:
The landing sites with SSR attachment sequences are pre-integrated into the genome with sufficient capacity to receive large transgene constructs. This preliminary preparation removes size constraints before transgene delivery, allowing integration of complex, large-scale genetic circuits and multiple genes simultaneously.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach increases the efficiency and accuracy of transgene integration, reduces the number of cell selections, and allows for rapid experimentation with various gene combinations, enhancing the versatility of genetic engineering.
Implementation Method 1
each landing site includes a pair of site-specific recombinase (SSR) attachment sites
Implementation Method 2
integrating a gene of interest into a landing site using prime editing
Data Source
AI summary
In variants, the method for generating a modular cell can include: integrating a landing site into a cell genome, and integrating a gene of interest into the landing site. The method can optionally include performing a cell selection, removing the gene of interest from the landing site, and/or any other suitable steps.