Gene Variant Libraries With Synonymous Codons for Accurate Activity Screening

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Solution Overview

Problem

Existing methods for testing protein variants are limited in accuracy and precision, particularly when distinguishing between variants that differ by a single nucleotide, leading to incorrect activity measurements due to sequencing errors.

Innovation Solution

Introducing synonymous codons into DNA encoding protein variants to increase the Hamming distance between variants, combined with compartmentalized self-replication (CSR) and shotgun sequencing, allows for precise measurement of the activity of thousands of variants.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If next-generation sequencing is used to measure variant activity, then high throughput measurement is achieved, but sequencing errors cause incorrect mapping between variants and functions

Engineering Contradiction:

Improvethroughput of variant activity measurementVSAvoidaccuracy of variant activity measurement

Core Design Contradiction:

ProductivityVSMeasurement precision

Solution Approach 1:

The patent applies preliminary action by introducing synonymous codons into the DNA library before sequencing to increase Hamming distance between variants. This pre-modification ensures that even if sequencing errors occur, the corrected sequences will still map to the correct variant, thereby resolving the contradiction between high throughput and measurement accuracy

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent implements beforehand cushioning by adding redundant synonymous codons that create a buffer against sequencing errors. The increased Hamming distance acts as a cushion that absorbs the impact of potential sequencing errors, ensuring accurate variant identification while maintaining high throughput measurement capabilities

Inventive Principle:

Principle #11Beforehand cushioning (Prior cushioning)

2Measurement precision

If variants are separated into different partitions for measurement, then activity of individual variants can be measured, but intermediate and low activity variants are not directly identified

Engineering Contradiction:

Improveability to measure individual variant activityVSAvoidinformation about intermediate and low activity variants

Core Design Contradiction:

Measurement precisionVSLoss of information

Solution Approach 1:

The patent uses copying by linking variant activity to the amount of DNA or RNA through PCR amplification and sequencing. This creates a quantitative copy of the activity information that can be measured, allowing detection of intermediate and low activity variants that would be missed in partition-based methods

Inventive Principle:

Principle #26Copying

3Device complexity

If Hamming distance between DNA variants is small (distance of one), then sequencing is faster and simpler, but sequencing errors result in incorrect mapping

Engineering Contradiction:

Improvesimplicity of sequencing processVSAvoidaccuracy of variant-function mapping

Core Design Contradiction:

Device complexityVSReliability

Solution Approach 1:

The patent applies parameter changes by deliberately increasing the Hamming distance parameter between DNA variants through the introduction of synonymous codons. This parameter modification maintains sequencing simplicity while dramatically improving the reliability of variant-function mapping by making sequences more resistant to sequencing errors

Inventive Principle:

Principle #35Parameter changes

Data Source

PatentUS20250340865A1Gene variant libraries and methods of use thereof

Publication Date: 2025.11.06 WATCHMAKER GENOMICS INC

AI summary

Disclosed herein, in part, are methods and compositions for performing high throughput screens of enzyme activity.