Disease Genes Detection Of Two Anterior Segment Abnormalities Pedigree By Whole Exome Sequence

Objective This study was to identify the disease-causing gene mutation in a family with ASD,and redicted the possible harm of the mutation by using the predictive software.Methods 1.Clinical examination: two ASD families comes from He Nan and He Bei provinces was enrolled in this study.The family members undergo a comprehensive clinical examination,including general examination and ophthalmic specialist examination.Eye specialist examinations include: best corrected visual acuity,examination using slit lamp,vitreous and fundus examination,fundus photography,ultrasound biomicroscopy,etc.2.Laboratory research: periphery blood specimens were collected from each family member ender the informed consent.The blood samples of 2 patients and 1 normal person in family 1 and 1 patient and 1 normal person in family 2 were send to the Novegene Bio Ptd to do the Whole Exome Sequences.The fragments of the mutation site were amplified in the whole family and digested with restriction endonuclease to confirm the relationship between the mutations and the patients.The candidate genes were verified by Sanger sequence and predicted damages by polyphen and SIFT software.Result: Family 1 included 17 members of 4 generations,and 9 patients were examined in serial 3 passages,which conformed to autosomal dominant inheritance pattern.Clinical examination revealed binocular congenital nuclear cataract in the 9 patients.All members in the family showed aniridia,choroidal lesions,and macular dysplasia.As the time goes by,patients II: 1,III: 1,III: 3 appear corneal opacity and secondary glaucoma,and complete loss sights in the end.And we found missense mutation T2A(p.M1K)on PAX6 gene;Family 2 including 8 members and 2patients were examined in serial 4 passages.Patients in this family showed aniridia,macular dysplasia,while the best corrected visual acuity can reach 0.1 to 0.2.We found splicing mutation c.357+1g>c on the same gene.Endonuclease digestion experiments confirmed that this site was heterozygous mutations.Missense mutation T2A(p.M1K)was gave a score of 0 in Polyphen-2,score of 0(harmful)in SIFT and Human splice founders software predicted two mutations that can affect m RNA cleavage and protein structure.Conclusions: T2A(p.M1K)and c.357+1g>c of PAX6 mutation are confirmed as pathogenic mutations of these 2 families respectively using whole exome sequencing and direct sequecing.The restriction endonuclease digestion test confirmed the mutation in family 1 as heterozygous mutation.Biological function prediction software Polyphen-2,SIFT and Human splicing founders predict the mutations will affect the prediction of m RNA cut and protein structure.Whole exome sequence provides a new approach to detect disease-causing mutations of congenital cataract with diversity clinical phenotypes.