Sequential DNA RNA Isolation Using Siliceous Supports
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Solution Overview
Problem
Current methods for isolating and purifying DNA and RNA from biological samples are challenging due to the complexity of natural biomaterials, often resulting in contamination and inefficiency, especially when using alcohols in wash buffers which can lead to alcohol contamination of the end product and hinder downstream processes.
Innovation Solution
A method involving sequential binding of DNA and RNA to different silicaceous materials using chaotropic salts and detergents, followed by enzymatic digestion and elution in alcohol-free buffers, allowing for the separation and purification of high-quality nucleic acids without alcohol contamination.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If alcohol-containing wash buffers are used to remove impurities from silicaceous matrices, then purification effectiveness is improved, but alcohol contamination of the eluted nucleic acid product occurs which hinders downstream processes
Solution Approach 1:
The invention extracts and removes alcohol from the wash buffer system entirely. Instead of using alcohol-containing buffers to wash impurities from nucleic acids bound to silicaceous matrices, the patent employs alternative washing strategies that achieve purification without introducing alcohol contamination that would interfere with downstream enzymatic reactions.
Solution Approach 2:
The invention introduces intermediate washing steps using alcohol-free buffers between the binding and elution stages. These intermediate buffers serve as mediators to remove impurities without the harmful alcohol component, allowing effective purification while maintaining compatibility with subsequent molecular biological procedures.
2Productivity
If DNA and RNA are isolated simultaneously from complex biological samples using conventional methods, then isolation efficiency is reduced due to competition for binding, but sequential isolation increases process complexity
Solution Approach 1:
The invention segments the nucleic acid isolation process into distinct sequential stages: first isolating DNA using conditions where DNA binds preferentially to the silicaceous matrix, then isolating RNA from the remaining sample under conditions where RNA binds preferentially. This segmentation eliminates competition between DNA and RNA for binding sites, improving the efficiency and purity of each isolated nucleic acid type.
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 isolates and purifies DNA and RNA from complex biological samples, ensuring high integrity and purity, with no RNA contamination in the DNA eluate and maintaining sample integrity, facilitating downstream molecular biological procedures.
Implementation Method 1
certain silicon-containing materials can absorb target substances in the presence of binding agents or binding enhancers
Implementation Method 2
The underlying mechanism for these nucleic acid isolation and purification methods is that silicon-containing materials can reversibly bind DNA, RNA and hybrid molecules of DNA and RNA in the presence of binding reagents. Some common chaotropic binding reagents include Nal, urea, guanidine hydrochloride, NaClO4, and KBr.
Implementation Method 3
Alcohol, such as 100% ethanol, is also a commonly used binding reagent for nucleic acid purification
Data Source
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Figure 2A~2D
Figure 3A~3C
AI summary
The invention provides a process and kit for serial isolation of DNA and RNA from the same sample. First, a siliceous solid support with preferential affinity for DNA over RNA is used to capture DNA in a lysate of a sample. Next, a siliceous solid support with similar affinity for RNA and DNA is used to capture RNA from the same lysate. The respective solid supports are recovered independent of each other, washed, and their bound nucleotide species are eluted. The invention further provides DNA and RNA prepared using the process in a minimal number of steps employing a minimal number of reagents. As the invention yields DNA and RNA of high quality and is amenable to automation, the invention may be used widely in the healthcare and pharmaceutical industries.