Plant Seed Heme Protein Expression Using Composite Regulatory Cassettes

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Solution Overview

Problem

Current methods fail to achieve high expression levels (>5%) of heme proteins in transgenic plant seeds, which are crucial for producing meat-like products with improved color, flavor, and nutritional value.

Innovation Solution

Utilizing a combination of seed-specific promoters (such as the 7S and beta-phaseolin promoters) and transcription/translation enhancers (like TEV and Rb7MAR) with optimized heme protein coding sequences, achieving expression levels of at least 5%, 8%, or 10% total soluble protein in transgenic plant seeds.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Quantity of substance

If conventional expression systems are used in transgenic plants, then the system complexity is low, but the expression level of heme proteins remains below 5% total soluble protein

Engineering Contradiction:

Improveexpression level of heme proteinVSAvoidcomplexity of expression cassette

Core Design Contradiction:

Quantity of substanceVSDevice complexity

Solution Approach 1:

The patent combines multiple regulatory elements (seed-specific promoters, transcription enhancers, translation enhancers, and polyadenylation signals) into a unified expression cassette to achieve high-level expression of heme proteins in plant seeds, resolving the contradiction between expression level and system complexity

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The expression cassette functions as a composite regulatory system integrating multiple DNA elements from different sources (plant promoters, viral enhancers, bacterial polyadenylation signals) to create a synergistic effect that achieves >5% heme protein expression in total soluble protein

Inventive Principle:

Principle #40Composite materials

2Quantity of substance

If seed-specific promoters and enhancers are used to increase heme protein expression, then the expression level exceeds 5% total soluble protein, but the regulatory elements required become more numerous and complex

Engineering Contradiction:

Improveexpression level of heme proteinVSAvoidnumber of regulatory elements

Core Design Contradiction:

Quantity of substanceVSDevice complexity

Solution Approach 1:

The expression cassette is segmented into distinct functional modules: seed-specific promoter region, transcription enhancer region, coding sequence, translation enhancer region, and polyadenylation signal region, allowing each element to be optimized independently while working synergistically to achieve high expression levels

Inventive Principle:

Principle #1Segmentation

3Reliability

If high expression levels of heme protein are achieved in plant seeds, then the nutritional value and color of plant-based products are improved, but the manufacturing process becomes more complex

Engineering Contradiction:

Improvenutritional value and color qualityVSAvoidease of producing transgenic plants

Core Design Contradiction:

ReliabilityVSEase of manufacture

Solution Approach 1:

The expression cassette is designed with all necessary regulatory elements pre-assembled in the correct configuration, allowing for straightforward transformation and expression of heme proteins in plant seeds without requiring multiple sequential genetic modifications, thereby simplifying the manufacturing process

Inventive Principle:

Principle #10Preliminary action

Data Source

PatentUS12359214B2High expression of animal heme protein in plants

Publication Date: 2025.07.15 MOOLEC SCIENCE LIMITED

AI summary

The present disclosure provides methods of producing heme proteins in transgenic plants, plant tissues, or plant cells, as well as describing the expression of these heme proteins in seeds. Also, the present disclosure provides transgenic plants expressing the heme proteins, myoglobin and hemoglobin, by introducing and integrating the recombinant DNA constructs into the host genetic material of the subject plants. The specific combination of regulatory elements disclosed herein allows for high heme protein expression level in seeds.