Multiplex KASP Primer Set for Wheat Height Gene Detection

Solution Overview

Problem

Current molecular marker-assisted selection methods for wheat plant height, particularly for Rht-B1 and Rht-D1 genes, are inefficient and cannot meet the demands of large-scale breeding due to limitations in throughput and cost, despite the potential of KASP markers for high-throughput SNP typing.

Innovation Solution

Development of a multiple KASP labeled primer set comprising specific primers for Rht-B1 and Rht-D1 genes, allowing simultaneous detection of different allelic variations through fluorescence detection, thereby improving detection efficiency and reducing costs.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If traditional PCR/electrophoresis markers are used for Rht-B1 and Rht-D1 genes, then detection can be performed, but throughput is limited to about several hundred samples daily and efficiency is low

Engineering Contradiction:

Improvedetection throughputVSAvoidtime consumption for detection

Core Design Contradiction:

ProductivityVSLoss of time

Solution Approach 1:

The patent combines detection of Rht-B1 and Rht-D1 genes into a single multiplex PCR reaction system, allowing simultaneous amplification and detection of both gene loci. This merging of detection processes enables throughput of more than 10,000 samples daily, dramatically improving productivity compared to traditional separate detection methods.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The patent replaces the mechanical electrophoresis separation system with a fluorescent detection system. By using KASP markers with fluorescently labeled primers and detecting allele-specific amplification through fluorescence intensity, the method eliminates the need for gel electrophoresis equipment and manual analysis, thereby increasing throughput and reducing time consumption.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

2Productivity

If traditional PCR/electrophoresis markers are used for Rht-B1 and Rht-D1 genes, then detection can be performed, but cost is high and efficiency is low

Engineering Contradiction:

Improvedetection efficiencyVSAvoiddetection cost

Core Design Contradiction:

ProductivityVSLoss of substance

Solution Approach 1:

The patent combines detection of Rht-B1 and Rht-D1 genes into a single multiplex PCR reaction system, allowing simultaneous amplification and detection of both gene loci. This merging of detection processes enables throughput of more than 10,000 samples daily, dramatically improving productivity compared to traditional separate detection methods.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The patent replaces the mechanical electrophoresis separation system with a fluorescent detection system. By using KASP markers with fluorescently labeled primers and detecting allele-specific amplification through fluorescence intensity, the method eliminates the need for gel electrophoresis equipment and manual analysis, thereby increasing throughput and reducing time consumption.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

3Measurement precision

If separate KASP markers are developed for Rht-B1 and Rht-D1 genes, then accurate detection is achieved, but detection time is doubled and cost increases

Engineering Contradiction:

Improvegene detection accuracyVSAvoiddetection throughput

Core Design Contradiction:

Measurement precisionVSProductivity

Solution Approach 1:

The patent combines detection of Rht-B1 and Rht-D1 genes into a single multiplex PCR reaction system, allowing simultaneous amplification and detection of both gene loci. This merging of detection processes enables throughput of more than 10,000 samples daily, dramatically improving productivity compared to traditional separate detection methods.

Inventive Principle:

Principle #5Merging (Combining)

4Measurement precision

If separate detection methods are used for Rht-B1 and Rht-D1 genes, then each gene can be accurately identified, but overall detection cost is doubled

Engineering Contradiction:

Improvegene identification accuracyVSAvoiddetection cost

Core Design Contradiction:

Measurement precisionVSLoss of substance

Solution Approach 1:

The patent combines detection of Rht-B1 and Rht-D1 genes into a single multiplex PCR reaction system, allowing simultaneous amplification and detection of both gene loci. This merging of detection processes enables throughput of more than 10,000 samples daily, dramatically improving productivity compared to traditional separate detection methods.

Inventive Principle:

Principle #5Merging (Combining)

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

The primer set enables simultaneous identification of Rht-B1 and Rht-D1 genes in a single PCR reaction, doubling efficiency and halving costs, making it suitable for large-scale wheat breeding applications.

Implementation Method 1

PCR amplification of wheat samples using the multiple KASP labeled primer set

Methodology Applied

Scientific EffectPCR amplification:

Implementation Method 2

The detection results can be directly obtained through fluorescence typing

Methodology Applied

Scientific EffectFluorescence detection: Fluorescence

Data Source

PatentUS12601014B2Multiple KASP marker primer set for wheat plant height major genes and use thereof

Publication Date: 2026.04.14 JIANGSU ACAD OF AGRI SCI

AI summary

A multiplex KASP marker primer set for a set of wheat plant height major genes, consisting of a pre-primer having a nucleotide sequence as shown in SEQ ID NO. 10, a post-primer having a nucleotide sequence as shown in SEQ ID NO. 11, and a universal primer having a nucleotide sequence as shown in SEQ ID NO. 9. The multiplex KASP marker primer set achieves the simultaneous detection of the Rht-B1 and Rht-D1 genes.