The invention discloses a second signal software calibration method of a clock chip, wherein the second signal software calibration method comprises the components of an RX-8025T clock chip, an HT6025 MCU chip, a clock calibration instrument, a microprocessor, an alarm circuit and an oscilloscope. The FOUT pin of the RX-8025T clock chip is electrically connected with an OSCI crystal oscillator input pin of the HT6025 MCU chip. An OSCO pin of the HT6025 MCU chip is suspended. The FOE pin of the RX-8025T clock chip is in short connection with the VDD pin. The FOUT pin of the HT6025 MCU chip is electrically connected with the clock calibration instrument. The microprocessor is electrically connected with the clock calibration instrument, the alarm circuit and the oscilloscope. The second signal software calibration method of the clock chip has advantages of high calibration precision, high calibration efficiency and effective production cost reduction.