A method for single-cycle selection of aptamers is provided. More specifically, a method comprising single-cycle selection of aptamers for proteins blotted on a membrane is provided. In some embodiments, the present methods can comprise a deoxyribonuclease I (DNase I) mediated aptamers selection strategy that may be capable of isolating strong binding aptamers for target proteins from a crude protein extract. Aptamers selected using the present method are further provided. More specifically, the presently selected aptamers may be thermally stable, modifiable and easily produced through single-cycle synthesis process. The present aptamers may provide unique affinity reagents for use in diagnosing and detecting infectious disease (for example Hepatitis B), for research and biochemical studies (e.g. into molecular mechanisms). Further, the present aptamers may be utilized to develop unique assays or kits for clinical application, such as monitoring disease treatment and outcome.