During the Upper Ordovician–Lower Silurian, chert was widely distributed in the Zhongbao Formation in the eastern part of the North Qilian Orogen. The origin and the tectonic setting of these chert were largely unknown. In order to analyse the material provenance, sedimentary environment, their formation and the tectonic setting, we present petrology and geochemical research on chert samples collected from Shihuigou Section. The evidence provided by radiolarite occurrences, Aluminium (Al)–iron (Fe)–manganese diagram and the silicon(Si)/Si + Al + Fe + calcium ratios suggesting a non-hydrothermal input and the biogenic origin chert. The geochemical features and the petrographic signatures have shown that the chert was also influenced by a terrigenous origin. It is considered that the deposition of the Late Ordovician chert is mainly affected by tectonic collision and volcanic ash events. During the Late Ordovician–Early Silurian transition, huge amounts of volcanic ash were released by massive volcanic activity that fell into the ocean, triggering the proliferation of radiolarians. Finally, in the Late Ordovician–Lower Silurian the tectonic setting of the North Qilian Orogen was not a typical deep-water basin, nor a typical continental margin, but a multi-island deep-water basin, which is closed to the mainland.