Two Novel Mutations In Fibrillin-1 Gene From Patients With Marfan Syndrome

Objective To screen mutation in the fibrillin-1 gene (FBN1)from 9 patients with Marfan syndrome (MFS) and identify the position and nature of FBN1 gene mutations. To observe the correlation between the clinical phenotype of patients with Marfan syndrome and the position or nature of FBN1 gene mutations.Methods Denaturing high-performance liquid chromatography (DHPLC) is introduced to screen for FBN1 mutation exon-by-exon. The DNA amplification fragments which DHPLC elution profiles show different from corresponding normal elution profiles are sequenced to identify the position and nature of mutation. The mutations can be further proved by allele-specific-PCR ( AS-PCR)or restriction-fragment-length-polymorphism (RFLP).Results DHPLC elution profiles of PCR products of exon34 from one patient (No.7) and that of exon43 from another MFS patient (No.6) are different from normal elution profiles. Direct sequencing showed a small insertion in exon34 at nucleotide 4307～4308 (4307insTCGT ) and a point mutation in exon43 at nucleotide 5309 (5309G>A). The insertion of 4 nucleotides results in frameshift mutation with premature termination codon and causes affected fibrillin-1 protein (Fib-1) shorter 1430 amino acids than normal Fib-1. The point mutation changes a highly conserved cysteine to tyrosine in cbEGF25(C1770Y).The two mutations are further confirmed by AS-PCR and RFLP respectively.Conclusions1. Frameshift mutation (4307insTCGT) in the exon34 and missense mutation (C1770Y) in the exon43 causes the corresponding patient to have MFS. 2. Mutation analysis of the FBN1 gene is helpful to identify individuals at high risk for MFS at an early stage, particularly in sporadic instances of the disease and in families of MFS displaying phenotypic variability, it also offers the possibility for a prenatal or preimplantation genetic diagnosis.3. DHPLC is a sensitive, convenient,automatic and highly-efficient method for mutation detection and can be widely used in mutation detection for MFS patients.