The invention provides a nucleic acid ligand for soft-shelled turtle iridovirus, and a construction method and application thereof. The nucleic acid ligand for the soft-shelled turtle iridovirus comprises a DNA sequence as shown in SEQ ID NO.1 and a derivative thereof. The nucleic acid ligand provided by the invention has relatively high affinity and specificity for the soft-shelled turtle iridovirus; and furthermore, the nucleic acid ligand also has the advantages of being free form immunogenicity, short in preparation period, good in reproducibility, small in molecular weight, convenient inin-vitro chemical synthesis, convenient to mark, easy to modify and substitute different parts of the nucleic acid ligand, steady in sequence, easy to transport and store, etc. When a rapid detectionmethod of the nucleic acid ligand in the invention is adopted for detecting the soft-shelled turtle iridovirus, operation is simple and rapid; the relatively high accuracy rate and sensitivity can beachieved; the nucleic acid ligand can be utilized for constructing a molecular probe or a detection kit; and furthermore, in combination with detection equipment, such as a microplate reader, a flow cytometry and a luorescence microscope, precise detection of the soft-shelled turtle iridovirus can be realized.