Purpose Relative anterior microphthalmos (RAM) is a rare congenital defect associated with severe vision impairment that is primarily caused by genetic alterations. The purpose of this study was to identify the causative genetic variants in two Chinese families with RAM with an autosomal recessive inheritance pattern. Methods DNA samples were obtained from two probands and their family members. Targeted next-generation sequencing (NGS) was used to screen 425 genes associated with inherited eye diseases to identify possible disease-causing variants in the two patients. Sanger sequencing was subsequently used to validate the results in both families. Results The targeted NGS panel identified potentially causative novel variants of the latent transforming growth factor beta binding protein 2 ( LTBP2) gene in the two RAM families: a missense variant (c.2771C > T; p.Ala924Val) and an intronic variant (c.4582 + 9A > G) in Family A and a different missense variant (c.5239C > A; p.Arg1747Ser) and a synonymous variant (c.951G > A; p.Pro317Pro) in Family B. These four novel variants all cosegregated with the disease phenotype. Conclusion To our knowledge, this is the first study to report novel LTBP2 gene variants related to RAM. Considering the importance of LTBP2 in ocular development, we provide initial insights into the potential pathogenic mechanisms of LTBP2 in RAM.