Selective Protein Removal from Nucleic Acid Samples
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Solution Overview
Problem
Current methods for purifying DNA from biological samples are inefficient as they often contaminate nucleic acids with proteins, as precipitating agents also precipitate nucleic acids along with proteins, requiring complex downstream processing to remove salts and other cellular components.
Innovation Solution
A method involving the addition of a specific compound, such as betaine, and a protein nucleating agent to a biological sample, followed by treatment with a hydrophobic surface material to selectively bind and remove proteins, allowing nucleic acids to remain in the solution.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If precipitating agents (e.g., TCA) are used to remove protein from biological samples, then protein removal efficiency is improved, but nucleic acids are also precipitated and lost
Solution Approach 1:
The invention modifies the chemical environment locally by introducing a specific compound that creates different precipitation conditions for proteins versus nucleic acids. This selective chemical modification allows proteins to precipitate while nucleic acids remain in solution, resolving the contradiction between protein removal efficiency and nucleic acid preservation
Solution Approach 2:
The invention changes the chemical parameters of the precipitation system by adding a specific compound that alters the precipitation behavior of biomolecules. This parameter change enables differential precipitation where proteins precipitate at conditions that do not cause nucleic acid precipitation, simultaneously achieving high protein removal efficiency while preventing nucleic acid loss
2Manufacturing precision
If conventional DNA purification methods (CsCl gradient, gel filtration, ion exchange chromatography) are used, then DNA purity is improved, but process complexity and time consumption increase
Solution Approach 1:
The invention extracts and removes the most problematic component (proteins) using a simplified selective precipitation method, eliminating the need for complex downstream purification steps. By taking out proteins through selective precipitation with the specific compound, the method achieves high DNA purity without requiring CsCl gradients, gel filtration, or ion exchange chromatography
Solution Approach 2:
The invention segments the purification process into distinct selective steps: first selectively precipitating proteins using the specific compound, then separately handling nucleic acids. This segmentation allows each component to be processed under optimized conditions, achieving high purity while reducing overall process complexity compared to conventional multi-step methods
3Manufacturing precision
If repeated alcohol precipitation and proteinase K treatment are used to purify plasmid DNA, then DNA purity is improved, but labor intensity and processing time increase
Solution Approach 1:
The invention performs preliminary selective protein precipitation using the specific compound before DNA isolation. This preliminary action removes proteins that would otherwise require time-consuming proteinase K treatment and repeated alcohol precipitations, significantly reducing total processing time while maintaining high plasmid DNA purity
Solution Approach 2:
The invention replaces mechanical and enzymatic processing steps (proteinase K treatment, repeated alcohol precipitations) with a chemical precipitation method using the specific compound. This substitution eliminates the need for labor-intensive manual operations and lengthy enzymatic incubations, reducing both labor intensity and processing time while achieving equivalent or superior DNA purity
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 effectively separates proteins from nucleic acids, enabling the use of the protein-free sample in subsequent processes like PCR without contaminating nucleic acids, thus simplifying DNA purification and reducing sample manipulation.
Implementation Method 1
It is known that trichloroacetic acid (TCA) can precipitate protein by three chloro groups contained therein
Implementation Method 2
nucleating agents that nucleate protein also precipitate nucleic acids, in addition to protein
Implementation Method 3
when a protein nucleating agent is added to a sample together with a specific compound, such as betaine, and the resultant mixture is treated with a hydrophobic surface material, protein can be efficiently removed
Data Source
AI summary
Provided is a method of removing protein while not removing nucleic acids from a biological sample containing protein, the method including: adding a compound of formula I below and a protein nucleating agent to the biological sample containing protein:where at least two of R1, R2, and R3 substituents are substituted or unsubstituted C1-C6 alkyl groups and the other substituent is a hydrogen atom or a substituted or unsubstituted C1-C6 alkyl group, a is an integer of 1 to 6, and b is 0 or 1, wherein b is 0 when a is not 1; treating the resultant mixture with a hydrophobic surface material in order to obtain a protein-free mixture; and separating the protein-free mixture from the hydrophobic surface material to which the protein is bound. By using the method, the protein can be selectively, effectively removed from the biological sample containing the protein while a nucleic acid is maintained in the sample.