Objective: To screen genes related to familial non-medullary thyroid carcinoma (FNMTC) using next-generation sequencing (NGS). Methods: A panel of NGS was designed and sequencing was performed for DNA samples extracted from peripheral blood leukocytes of FNMTC patients and sporadic non-medullary thyroid carcinoma (SNMTC) cases, respectively, and gene mutations were screened. In addition, the clinicopathological characteristics, including tumor size, extension of surgery, lymph node metastasis and extra-thyroidal extension, were compared between patients with or without mutations. Results: In 63 NMTC samples, 45 mutations were detected on 13 genes. 37 germline mutations were detected in 47 FNMTC patients, while 8 germline mutations were detected in 16 SNMTC patients. In 8 FNMTC family lineages, the same mutations were carried by FNMTC patients from the same pedigree. The number of carriers of mutations was 29 in the 47 FNMTC patients and 6 in the 16 SNMTC patients, with a non-significant difference (P= 0.092). Among the FNMTC patients, there were 22 patients with central lymph node metastasis in the 29 mutation-positive patients, significantly more than 7 in the 16 mutation-negative cases (P= 0.031). As for the parentage, there were 3 patients with central lymph node involvement among the 7 patients of parent generation, while all the 9 patients of offspring generation had central lymph node metastasis (P=0.019). Conclusions: This panel of NGS can be used to screen mutant susceptibility gene of FNMTC patients, and the findings may be helpful for early detection of FNMTC patients and predicting the disease risk to familial members of FNMTC patients.