Aim To generate mutant mouse with ALK7 gene inserted by LoxP sequences by CRISPR/Cas9, to further develop temperospatial specific ALK7 deleted mouse models via cross bred with the tissue-specific Cre mouse, providing a basis for functional study of ALK7 in special time and tissue. Methods The CRISPR/Cas9 technology was used to edit ALK7. Two sgRNAs were designed to direct Cas9 endonuclease cleavage in intron 3-4 and intron 6-7. The targeting vector with LoxP-ALK7-LoxP sequence was designed. sgRNA, Cas9 mRNA and targeting vector were co-injected into zygotes, which were transferred to pseudopregnant mice. The pups were genotyped by PCR and Southern blott. Real-time PCR and Western blot were performed for analysis of the expression of ALK7. Results The ALK7LoxP/LoxP mouse was generated by CRISPR/Cas9, and did not show influence on the expression of ALK7. Conclusion The CRISPR/Cas9 technology can successfully generate the ALK7LoxP/LoxP mouse by inserting LoxP sequence into ALK7 gene, which is a basis for the creation of tissue-specific ALK7 deleted mouse models.