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Updated: May 11, 2026
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Primer-Free Aptamer Selection Using A Random DNA Library
Published on: July 26, 2010
Single-primer-limited amplification: a method to generate random single-stranded DNA sub-library for aptamer
Chao-Zhu He1, Kun-He Zhang, Ting Wang
1Department of Gastroenterology, First Affiliated Hospital of Nanchang University, Jiangxi Institute of Gastroenterology and Hepatology, Nanchang 330006, China.
Analytical Biochemistry
|May 29, 2013
Summary
Single-Primer-Limited Amplification (SPLA) efficiently generates DNA libraries for aptamer selection. This method minimizes by-products, improving DNA amplification for systematic evolution of ligands by exponential enrichment (SELEX).
Area of Science:
- Biotechnology
- Molecular Biology
- Genomics
Background:
- Polymerase chain reaction (PCR) amplification of single-stranded DNA (ssDNA) libraries is crucial for aptamer selection.
- Nonspecific hybridization during PCR can lead to by-product amplification, hindering aptamer development.
- Existing methods struggle to produce clean ssDNA libraries for systematic evolution of ligands by exponential enrichment (SELEX).
Purpose of the Study:
- To develop a novel method for amplifying random ssDNA libraries free from by-products.
- To optimize conditions for efficient and specific ssDNA amplification for aptamer selection.
Main Methods:
- Developed Single-Primer-Limited Amplification (SPLA) using limited primer quantities.
- Initiated amplification with limited reverse primer for minus-stranded DNA (msDNA).
- Followed with limited forward primer for plus-stranded DNA (psDNA) and purified via gel excision.
Main Results:
- SPLA effectively amplified target ssDNA without detectable by-products or nonspecific products.
- Optimized SPLA produced 16.1 times more psDNA compared to asymmetric PCR.
- By-product formation correlated with increased template amount and thermal cycles.
Conclusions:
- SPLA is a simple and feasible method for generating high-quality random ssDNA sub-libraries.
- This technique significantly improves the efficiency of aptamer selection processes.
- SPLA offers a robust alternative to conventional PCR for ssDNA library amplification.
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