Super Recombinator System for Multi-Transgene Integration
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Solution Overview
Problem
Traditional methods for creating organisms with multiple transgenic elements are inefficient due to genetic linkage and crossing-over constraints, requiring numerous steps and combinations, making it difficult to integrate multiple transgenes at the same locus.
Innovation Solution
The Super Recombinator (SuRe) system uses adaptor sequences and linker sequences to facilitate recombination, reducing the number of steps required to integrate multiple transgenic elements at the same locus from N−1 to log2N steps and simplifying the selection process by using orthogonal linkers and markers.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional recombination methods are used to integrate multiple transgenes, then the number of recombination steps increases linearly (N-1 steps for N transgenes), but the efficiency and time required for creating multi-transgenic organisms decreases
Solution Approach 1:
The patent introduces adaptor sequences as intermediary elements that facilitate recombination between transgenes. These adaptors contain specific linker sequences that enable controlled recombination events, reducing the number of steps required to integrate multiple transgenes at the same locus. The adaptors act as mediators that bridge transgenes together through homologous recombination, significantly improving recombination efficiency compared to traditional methods.
Solution Approach 2:
The patent segments the recombination process into modular steps using standardized adaptor sequences with specific linker regions. By dividing the complex multi-transgene integration into manageable recombination events between adjacent adaptors, the system achieves logarithmic scaling (log2N steps) rather than linear scaling. Each adaptor-transgene unit can be independently manipulated and combined.
2Adaptability or versatility
If multiple transgenes are integrated at the same locus using traditional methods, then the number of possible genetic combinations increases exponentially (2^N+1 in diploids, 4^N+1 in tetraploids), but the difficulty of selecting correct recombinants increases
Solution Approach 1:
The patent applies local quality by making each adaptor sequence unique with specific linker sequences that only recombine with their complementary partners. This localized specificity at each recombination site prevents unwanted recombination events between non-paired adaptors, even when multiple transgenes are present. The orthogonal nature of different adaptor pairs ensures that recombination occurs only at intended locations, simplifying selection of correct recombinants.
Solution Approach 2:
The adaptor sequences serve as intermediaries that carry specific genetic markers and linker sequences. These intermediaries facilitate controlled recombination by providing defined recombination sites with unique sequences. The markers within adaptors enable straightforward identification of correct recombinants, reducing the complexity of selection processes even as the number of transgenes increases.
3Quantity of substance
If traditional docking sites are used for transgene integration, then the number of available docking sites is limited, but the number of transgenic elements desired to be integrated exceeds the available sites
Solution Approach 1:
The patent creates a universal recombination system where adaptor sequences with standardized linker regions can be used with any transgene. This universal approach allows multiple transgenes to be integrated at the same genomic locus through sequential recombination events. The standardized adaptor design enables the system to handle an unlimited number of transgenic elements, far exceeding the limitations of traditional fixed docking sites.
Solution Approach 2:
The patent transitions from integrating transgenes at different genomic locations (spatial dimension) to integrating multiple transgenes at the same locus through temporal sequencing of recombination events. By using adaptor sequences that enable controlled recombination at a single site, the system effectively adds a temporal dimension to the integration process, allowing N transgenes to be integrated at one locus through log2N sequential steps rather than requiring N different loci.
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 significantly increases the efficiency of transgene recombination, reducing labor and time, and allows for the creation of organisms with multiple transgenic elements in a more streamlined and cost-effective manner, as demonstrated by higher recombination efficiencies compared to natural recombination methods.
Implementation Method 1
facilitating recombination between a matching linker sequence within each of the first adaptor and the second adaptor
Data Source
AI summary
Provided herein are materials and methods useful for facilitating transgene recombination. The present disclosure relates to, for example, techniques for manipulating recombination frequencies and generating organisms that contain multiple transgenic elements docking at the same locus on a single chromosome. The time consumed by the entire recombination process is proportional to the logarithm of the number of transgenes to be recombined.