Plant Nucleic Acid Isolation Using Non-Spherical Disrupting Particles
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Solution Overview
Problem
Nucleic acid isolation from plants is challenging due to the difficult-to-lyse nature of plant cells and the presence of inhibitors like polysaccharides and polyphenolic compounds, leading to low yields and poor quality DNA, especially when using methods like mortar and pestle or mechanical disruption with common grinding media.
Innovation Solution
A method involving mechanical disruption of plant samples using a liquid lysis composition with chaotropic agents and non-spherical solid disrupting particles, followed by contacting the lysate with protein precipitating and inhibitor removing agents to isolate nucleic acids.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If mortar and pestle or mechanical disruption with common grinding media is used, then plant cell lysis can be achieved, but nucleic acid yield is low and inhibitor removal is insufficient
Solution Approach 1:
The patent changes the physical and chemical parameters of the lysis system by using non-spherical solid disrupting particles instead of conventional spherical beads or mortar and pestle. The non-spherical particles create more effective mechanical disruption forces that efficiently lyse plant cells while releasing nucleic acids. Additionally, the patent modifies the chemical environment using specific lysis buffers containing chaotropic agents and inhibitors to optimize both lysis efficiency and inhibitor removal, thereby achieving high nucleic acid yield with minimal contamination.
Solution Approach 2:
The patent introduces a specialized lysis buffer system as an intermediary medium that facilitates both cell lysis and inhibitor removal. This buffer contains chaotropic agents that disrupt cellular structures while protecting nucleic acids, and inhibitor removal agents that specifically target polysaccharides and polyphenolic compounds. The intermediary buffer system enables simultaneous achievement of effective lysis and purification without requiring separate steps.
2Productivity
If CTAB lysis buffer is used for lysis and inhibitor removal, then nucleic acid isolation can be performed, but CTAB is toxic and the method is time-consuming
Solution Approach 1:
The patent replaces CTAB with alternative chaotropic agents and inhibitor removal agents that are less toxic while maintaining effective lysis and purification functions. The new chemical system achieves the same nucleic acid isolation efficiency without the harmful effects of CTAB, thereby improving safety while preserving productivity.
3Adaptability or versatility
If standard lysis methods are used, then plant material can be processed, but the protocol cannot be universally applied to different plant types
Solution Approach 1:
The patent develops a universal lysis protocol using non-spherical solid disrupting particles and a comprehensive lysis buffer system that can effectively process multiple plant types. The non-spherical particles provide consistent mechanical disruption across different tissue structures, while the buffer system contains broad-spectrum inhibitors that work on various plant species. This multi-functional approach enables high-quality nucleic acid isolation from diverse plant samples using a single standardized protocol.
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 method achieves high-quality nucleic acid isolation with high yield from various plant samples by efficiently lysing plant material and removing inhibitors, providing a universal protocol for different plant types.
Implementation Method 1
liquid lysis composition which comprises at least one chaotropic agent
Implementation Method 2
mechanically disrupting the plant sample in a liquid lysis composition which comprises at least one chaotropic agent and one or more solid disrupting particles
Implementation Method 3
contacting the lysed sample with at least one protein precipitating agent
Implementation Method 4
contacting the lysed sample with at least one inhibitor removing agent
Data Source
AI summary
Provided is a method for isolating nucleic acids from a plant sample comprising(a) preparing a lysed sample wherein preparing comprises(i) lysing a plant sample by mechanically disrupting the plant sample in a lysis solution which comprises at least one chaotropic agent and one or more solid disrupting particles, and(ii) optionally clearing the lysate;(b) contacting the lysed sample with at least one protein precipitating agent and at least one inhibitor removing agent and providing a mixture;(c) obtaining a liquid phase from the mixture; and(d) isolating nucleic acids from the liquid phase.Also provided is a kit for use in such method.