In this work, we systematically study the transmission spectral characteristics of the microfiber knot resonator (MKR) both in simulation and in an experiment. The transmission spectrum shows several possible phenomena, including dominating resonance, dominating interference, the coexistence of resonance and interference with different intensity ratios, and resonance with Vernier effect. The transmission spectra with various shapes are obtained in simulation by adjusting parameters like fractional coupling intensity loss, intensity coupling ratio, and propagation constants of different modes. The relationship between the transmission spectrum and the size of the resonant ring is studied by reducing the diameter of the resonant ring and analyzing the corresponding transmission spectrum. The performance of a polydimethylsiloxane-coated MKR in refractive index and temperature measurement is also investigated, and the responses of the transmission spectrum, the extracted interference spectrum, the extracted resonance spectrum, the extracted envelope spectrum of Vernier effect are all analyzed, indicating that these spectra have similar responses to the variation of refractive index or temperature. Linear fit indicates that the sensitivities of the transmission spectrum to refractive index and temperature are about 1264.09 nm/RIU and -2.60 nm/℃, respectively. This systematic and comprehensive study will provide a helpful guideline for understanding, analyzing and designing MKR-based devices with desired spectral characteristics to meet different application requirements.