A key operation in modern genetic engineering is to deliver foreign substances (like modified subcellular organelles, DNA molecules, and drugs) into live cells. Compared with the traditional methods like polyethyleneimine (PEI), cationic liposomes, viral vectors, electroporation, gene gun, etc., microinjection is a rising star due to the advantages of excellent safety, full controllability, high precision, and high survival rate. This review focuses on the recent development of microinjection technology and its applications in animal cells, yeast, fungal cells, and plant cells. Guidelines are provided for selecting the most appropriate microinjection method and system configuration for different application scenarios. When the outer membrane of the host cell is difficult to break, the piezo-driven penetrator could assist the penetration significantly and causes less damage to the host cell during microinjection process. The major existing challenges in microinjection are also analyzed. Overall, microinjection could become a standard technology for modern genetic engineering and microbiology.