Construction Of Targeting Vector For Knockin Of PRPS1and Mutational Screening Of TCOF1Gene In Patients With Treacher Collins Syndrome

Deafness is common disease led to obstacles of communication, caused by geneticand environmental factors.60%of deafness patients are are hereditary,which ismainly a monogenic genetic disease. In china, with the rapid development ofeconomy and the improvement of our living standards, the effects of environmentalfactors to deafness are decreased gradually.The importance of genetic factors in thecause of deafness is increasingly prominent. In recent years, with the rapiddevelopment of molecular genetics, our understanding of the auditory function andhearing loss causes is getting more and more in-depth. Some important researchfindings have been applied in clinical practice for genetic diagnosis of deafness. Butsome gene functions and mechanism lead to deafness still remain unclear. Morein-depth research on the etiology and pathogenesis of hereditary deafness is needed tolook for a forecast, reducing the risk of offspring deafness and new treatment methods,and improving the quality of our population, which has important practical andlong-term significance. In this study, the etiology and pathogenesis of hereditarydeafness were studied, which is divided into two parts:PART1: Construction of targeting vector for knockin of PRPS1.In the previous study, our group has reported a novel gene, PRPS1, underlyinghuman DFNX1deafness, and has deciphered its mechanism. By collaborating withseveral international groups, we have identified different PRPS1mutations in otherDFNX1families, which proved our finding. From the functional study of DFNX1mechanism, we found that using SAM has the potential to prevent DFNX1presymptomly (PRPS1is a key enzyme in pathway of purine and pyrimidinesynthesis). A mouse model mimicking human DFNX1deafness is needed to test thevalidity and security of SAM in DFNX1prevention in human. In this study, we haveestablished a gene vector for knockin of PRPS1by Gene targeting technology.Targeting ES cells from129strain mice by electro-transformation. The targeted EScells were screened out by antibiotic G418and PCR reaction. This vector provides abasis for establishment of DFNX1mouse model, PRPS1gene function research, prevention before onset, and therapeutic approaches.PART2: Mutational screening of TCOF1gene in patients with TreacherCollins syndromeTreacher Collins syndrome (Treacher Collins syndrome, TCS) is also known asFranceschetti syndrome, Franceschetti-Zwahlen-Klein Syndrome, is an autosomaldominant disease affecting the facial development, also known as the mandibularbone hypoplasia (Mandibulofacial Dysostosis, MFD),The typical physical featuresinclude downward slanting eyes, micrognathia, underdevelopedzygoma, conductivehearing loss, drooping part of the lateral lower eyelids, and malformed or absent ears.We have collected one sporadic patients with conductive hearing loss and notching ofthe lower eyelid recruited from Outpatient Department of our hospital. By directsequencing TCOF1, we identified a novel missense mutation c.1639delAG in the11thexon of TCOF1gene in patient, this mutation have a termination codon (p.S547X),but was not found in the gene in his parents. Results of this study can be used toconduct prenatal diagnosis in this family.