The invention relates to a high-efficient culture and sorting method and a culture device of microorganisms. The culture device is characterized in that the culture device comprises a temperature-controlled culture box, a plurality of culture medium bottles are arranged on an upper shelf in the temperature-controlled culture box, the corresponding number of chromatography columns are arranged on a lower shelf, a hose is arranged therebetween for communication, a flow rate controller is arranged on the hose, silk sieve cloth with 25mu m is covered on an outlet end of each chromatography column, and the access into a waste liquid collection bottle is realized through the hose. The high-efficient culture and sorting method is as follows: 1) collection and concentration of the microorganisms;2) embedding of the microorganisms; 3) culture of the embedded microorganism under near-nature conditions; and 4) sorting and enrichment culture of the microorganisms with monoclonal microspheres. The method can carry out high-efficient culture and sorting of the microorganisms, provide a new path for obtaining the pure culture of the microorganisms, in particular to new species of the microorganisms, greatly reduce the workload of obtaining the pure culture of the microorganisms by using the traditional method and lay a foundation for 16rDNA sequencing of the microorganisms, determining the species thereof and studying gene functions and metabolic products of the microorganisms, and other downstream studies.