Nucleic Acid Nanoball Condensation for Uniform DNB Loading
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Solution Overview
Problem
Existing nucleic acid sequencing technologies face challenges in achieving high sequencing quality due to increased DNB copy number leading to larger volumes, which cause mutual interference and non-uniform sizes during loading, resulting in empty sites and deteriorated sequencing quality.
Innovation Solution
A method involving condensing treatments with nucleic acid condensing agents and adjusting pH to 3-4.5 to compress and uniformize the volume of nucleic acid nanoballs, using agents like multivalent salts, basic peptides, and polar solvents, to improve loading efficiency and quality.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Illumination intensity
If the RCA time is extended to increase the DNB copy number, then the detection signal becomes stronger, but the DNB volume increases leading to greater mutual interference during loading
Solution Approach 1:
The patent applies parameter changes by adjusting the pH of the loading buffer to 3-4.5, which alters the physical state of the DNBs. This pH adjustment causes the DNBs to transition from a loose, expanded state to a condensed, compact state, thereby reducing their effective volume while preserving the high copy number achieved through extended RCA time. The condensed state eliminates mutual interference during loading while maintaining the signal strength benefits of high copy numbers.
2Reliability
If the DNB copy number is increased to strengthen the detection signal, then sequencing quality improves, but the DNB size becomes larger causing non-uniform distribution on the chip
Solution Approach 1:
The patent utilizes parameter changes by controlling the pH of the loading buffer within the range of 3-4.5. This pH parameter adjustment induces a conformational change in the DNB structure, causing them to adopt a condensed state with uniform size and shape. This ensures that even with increased copy numbers for enhanced sequencing quality, the DNBs maintain consistent dimensions that enable precise and uniform loading onto the chip array.
3Measurement precision
If the DNB copy number is increased to enhance signal strength, then base calling accuracy improves, but empty sites increase due to insufficient reactions from mutual interference
Solution Approach 1:
The patent applies parameter changes by adjusting the loading buffer pH to 3-4.5, which condenses the DNB volume and reduces their hydrodynamic radius. This condensation effect allows a higher density of DNBs to be loaded onto the chip without mutual interference, thereby increasing loading efficiency and reducing empty sites. Simultaneously, the high copy number within each condensed DNB maintains strong detection signals for accurate base calling.
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
The method results in more uniform distribution and stronger signal concentration of nucleic acid nanoballs on solid supports, reducing empty sites and enhancing sequencing quality, particularly for long-read sequencing.
Implementation Method 1
adding a nucleic acid condensing agent to a system comprising the nucleic acid nanoball
Implementation Method 2
adjusting a pH of the system comprising the nucleic acid nanoball to 3-4.5
Data Source
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AI summary
A method for adjusting the volume of nucleic acid nanoballs. By performing coagulation treatment on a nucleic acid (for example, adding a nucleic acid coagulating agent and/or adjusting the pH value to 3-4.5, so that positively charged counter ions are gathered around the nucleic acid, and negative charges on the surface of the nucleic acid are neutralized by cations by means of electrostatic bonding of anions and cations, thereby enabling the nucleic acid to coagulate), the shape of the nucleic acid nanoballs can be changed, the volume of the nucleic acid nanoballs can be adjusted, and the nucleic acid nanoballs can be compressed. The nucleic acid nanoballs obtained by the method (performing coagulation treatment on the nucleic acid nanoballs) are more compact, smaller in shape, and more uniform in size. The method can be used in any situation where the volume of the nucleic acid nanoballs needs to be adjusted (reduced) (such as the loading process of nucleic acid nanoballs (such as DNBs), and the sequencing process after loading).