The invention discloses a fully-closed pollution-free digital nucleic acid detection chip which comprises a chip body, a first puncture needle and a second puncture needle, the chip body is made of a transparent material, a plurality of oil inlet through holes are formed in the chip body, outlets of the oil inlet through holes are connected with an oil inlet groove, one end, connected with the first puncture needle, of the oil inlet groove is provided with an opening, an opening in the other end of the first puncture needle extends into a test tube, the test tube is used for receiving an oil phase conveyed by the oil inlet through holes, an opening in one end of the second puncture needle is connected into a liquid inlet groove formed in an inner cavity of the chip body, an outlet of the liquid inlet groove is connected with a tiling cavity, the bottom plane of the tiling cavity is in the horizontal direction, and an opening in the other end of the second puncture needle extends into the test tube; and the opening in the other end of the first puncture needle extends into the oil phase in the test tube all the time, and when the test tube is filled with the oil phase, the opening in the other end of the second puncture needle is in contact with the top liquid level of the oil phase. The risk of cross contamination is avoided, operation is easy, and consumed time is short.