NLR Gene Library Preparation for Crop Disease Resistance

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Solution Overview

Problem

Current methods for introducing multiple plant disease resistance genes into crop plants are laborious and time-consuming, often involving linkage drag and requiring the production of susceptible plants for mutagenesis, which can be challenging, and traditional map-based cloning is inefficient due to the complexity of nucleotide-binding leucine-rich repeat (NLR) gene clusters in plant genomes.

Innovation Solution

A method for preparing a library of candidate NLR genes by selecting highly expressed NLRs from unchallenged plant tissues, followed by transforming host plants with these genes and testing for resistance to specific plant pathogens, allowing for the rapid identification and deployment of effective resistance genes without the need for susceptible plant mutagenesis or extensive map-based genetics.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If traditional breeding methods are used to deploy multiple R genes, then the genes can be introduced into the crop plant, but the process becomes extremely laborious and time consuming due to the scattered distribution of R genes throughout the genome

Engineering Contradiction:

Improvespeed of deploying multiple R genesVSAvoidtime required for combining multiple R genes

Core Design Contradiction:

ProductivityVSLoss of time

Solution Approach 1:

The patent merges multiple R genes into a single integrated cassette that functions as one genetic unit. This cassette can be simultaneously introduced into a crop plant through a single transgenic event, eliminating the need for separate breeding operations to combine multiple scattered R genes throughout the genome.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The patent uses a synthetic promoter and common intron as intermediary elements that facilitate the expression of multiple R genes within the cassette. These intermediary components ensure that all R genes in the cassette are properly expressed together, enabling functional deployment without complex breeding procedures.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Measurement precision

If traditional map-based cloning is used to identify R genes, then the genes can be isolated, but the process remains challenging and inefficient due to the complexity of NLR gene clusters and lack of recombination in large genomic tracts

Engineering Contradiction:

Improveidentification of specific R genesVSAvoidcomplexity of NLR gene clusters

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent extracts and sequences the coding regions of R genes from wild relative genomes using RNA-seq data, separating the identification process from the complex genomic context. This allows precise identification of R genes without being hindered by the lack of recombination or the complexity of NLR clusters in the original genome.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent creates synthetic copies of R genes from wild relatives and introduces them into the crop plant genome. This copying approach allows the R genes to be deployed without needing to isolate them through complex map-based cloning procedures, bypassing the limitations of working with complex NLR clusters.

Inventive Principle:

Principle #26Copying

3Reliability

If R genes are deployed one at a time, then the genes can be introduced into the crop plant, but the pathogen overcomes the resistance within a few seasons

Engineering Contradiction:

Improveduration of disease resistanceVSAvoidrate of resistance deployment

Core Design Contradiction:

ReliabilityVSProductivity

Solution Approach 1:

The patent combines multiple R genes into a single cassette that is introduced together into the crop plant. This simultaneous deployment of multiple R genes creates a composite resistance system that is more durable against pathogens, as the pathogen would need to overcome multiple resistance mechanisms at once rather than sequentially.

Inventive Principle:

Principle #5Merging (Combining)

4Adaptability or versatility

If multiple R genes are scattered throughout the plant genome, then the genes can be present in the plant, but combining them into a single plant becomes extremely laborious

Engineering Contradiction:

Improvepresence of multiple R genesVSAvoidease of combining R genes

Core Design Contradiction:

Adaptability or versatilityVSEase of manufacture

Solution Approach 1:

The patent merges multiple R genes into a single integrated cassette structure. This allows the plant to possess multiple R genes while making their combination straightforward, as all genes are delivered together in one transgenic event rather than requiring separate breeding operations to assemble scattered genes.

Inventive Principle:

Principle #5Merging (Combining)

Data Source

PatentUS20240218356A1Methods for preparing a library of plant disease resistance genes for functional testing for disease resistance

Publication Date: 2024.07.04 TWO BLADES FOUND

AI summary

Methods are provided for preparing a library of candidate plant disease resistance (R) genes against a plant pathogen of interest. The methods involve selecting from each of one or more plants of interest a subpopulation of highly expressed nucleotide-binding leucine rich repeat genes (NLRs) from among the population of NLRs that are constitutively expressed in an organ or other part of the one or more plants to produce a library of candidate R genes. Further provided are related methods for identifying R genes against a plant pathogen of interest using a library of candidate R genes and compositions comprising the identified R genes.