The invention discloses a method for extracting wolbachia used for embryonic injection transfection, which comprises: dissecting an insect having wolbachia to obtain the ovary of the insect, and placing the ovary in sucrose phosphate glutamate (SPG) buffer solution; and homogenizing the ovary, washing the homogenate with the SPG buffer solution, collecting the homogenate, placing the collected homogenate in a centrifuge to centrifuge the homogenate in amount of 100 to 1,000 grams for 1 to 10 minutes, collecting supernate, centrifuging the supernate in an amount of 5,000 to 15,000 grams for 5 to 20 minutes, removing supernate, washing the sediment with the SPG buffer solution to obtain suspension, centrifuging the suspension in an amount of 100 to 1,000 grams for 1 to 10 minutes, collecting the supernate which is pure wolbachia, wherein the pure wolbachia can be used for embryonic injection in mosquitoes. In the method disclosed by the invention, the source (ovary tissues of insects having the wolbachia) of the wolbachia is rich and can be obtained easily; and the whole method is simple and convenient in operation, has very high controllability, and can obtain lots of wolbachia forembryonic injection transfection in a give time very easily, thereby saving lots time and energy, improving working efficiency and contributing to quick establishment of a mosquito strain transfectedwith wolbachia.