Total DNA Isolation from Bodily Fluids via Protein Digestion
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Solution Overview
Problem
Current methods for isolating DNA from bodily fluids, such as urine, are invasive, inefficient, and primarily isolate only free DNA, resulting in low yields and loss of protein-bound DNA, necessitating large sample volumes for sufficient DNA extraction.
Innovation Solution
A method involving the concentration and isolation of both free DNA and proteins, followed by protein digestion to release protein-associated DNA, using silicon carbide columns or slurry formats, allowing for the purification of total DNA from small bodily fluid samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If classical methods are used to isolate DNA from urine, then free DNA can be isolated, but protein-bound DNA is lost and yield is low
Solution Approach 1:
The isolation process is divided into distinct segments: first isolating proteins with bound DNA, then separately isolating free DNA, and finally combining both fractions. This segmentation allows recovery of previously lost protein-bound DNA while maintaining free DNA isolation efficiency
Solution Approach 2:
Proteins with bound DNA are isolated first as a preliminary step before free DNA isolation. This preliminary action prevents loss of protein-bound DNA and enables subsequent release of bound DNA through protein digestion, increasing total DNA yield
2Quantity of substance
If large volumes of urine are processed, then sufficient DNA can be obtained, but the process becomes tedious and time-consuming
Solution Approach 1:
The method changes the isolation parameters by using specific buffers and conditions that enable efficient DNA binding to silica-based columns from smaller urine volumes. This allows sufficient DNA recovery from reduced sample volumes without extending processing time
Solution Approach 2:
The method extracts and concentrates DNA from small urine volumes using silica-based columns that selectively bind DNA. This extraction approach obtains sufficient DNA quantity from minimal sample volume, eliminating the need to process large volumes and reducing overall isolation time
3Quantity of substance
If invasive methods like amniocentesis or biopsy are used, then DNA can be isolated for diagnostics, but patient safety risks and pain increase
Solution Approach 1:
The method utilizes DNA that is naturally present and shed into urine from the urinary tract and circulation. This self-service approach eliminates the need for invasive procedures by harnessing the body's natural DNA release mechanisms, providing diagnostic DNA without patient risk or pain
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 approach enables the isolation of larger quantities of biologically active total DNA from small volumes of bodily fluids, including urine, serum, and other samples, enhancing diagnostic applications by recovering both free and protein-bound DNA.
Implementation Method 1
The free DNA and proteins are bound to a resin, such as silicon carbide particles
Implementation Method 2
The proteins can then be digested using a range of enzymes, including proteinase K and pronase
Implementation Method 3
The released DNA can then be isolated by precipitation, binding it to a resin that is either packed in a column or in a slurry format
Data Source
AI summary
A method is disclosed for isolating both free and protein-associated DNA from bodily fluids, such as urine, saliva, serum, tears, sweat, cerebral spinal fluid, and plasma. The method comprises as a first step concentrating and isolating both the free DNA and the proteins present in the bodily fluid. The proteins are then digested in order to release the formerly protein-associated DNA from the isolated proteins. Lastly, the free and formerly protein-associated DNA can be isolated and purified.