Mutational analysis of the rhodopsin gene in Chinese ADRP families by conformation sensitive gel electrophoresis.

Retinitis pigmentosa is a very heterogeneous group of retinal degenerations, with multiple genes identified in each mode of inheritance. For autosomal dominant retinitis pigmentosa (ADRP), the most common gene is the rhodopsin (RHO) gene, mutations in which contribute to about 25% of ADRP in Caucasian population. To investigate the frequency and pattern of RHO point mutations in Chinese patients with ADRP, we have screened the five coding exons and splice sites of the RHO gene in 50 unrelated probands from Chinese ADRP families and 100 normal controls to identify disease-associated mutations, using conformation sensitive gel electrophoresis (CSGE) and direct DNA sequencing. Two RHO mutations, Pro347Leu and Pro327 (1-bp del), were identified each in one family, thus the frequency of RHO mutations among ADRP families in this study is less than 14% (2/50=4%, 95% confidence interval: 1-14%), lower than that in Europe and North America, which may reflect an ethnic difference between Chinese and Caucasian populations. Loss of all phosphorylation sites at the C-terminus and a highly conserved sequence QVS(A)PA may occur because of Pro327(1-bp del). CSGE was found to be a sensitive, simple and practical method for the screening of a large number of samples under highly reproducible conditions, and could be utilized in routine molecular diagnostic laboratories.

Molecular studies of loss of heterozygosity in Chinese sporadic retinoblastoma patients.

BACKGROUND: Retinoblastoma, an embryonic neoplasm of retinal origin, is the most common and severe intra-ocular tumor affecting infants and young children. METHODS: Loss of heterozygosity (LOH) on chromosome 13 was investigated in 16 Chinese sporadic RB patients, using 14 microsatellite markers spanning the complete chromosome 13, to determine whether those alterations different from the alteration of RB1 gene on 13q14 may play a role in the development of RB. Loss of RB1 allele is commonly encountered in sporadic RB. Microdissected RB tissues and their matched blood DNAs were analyzed for PCR-based LOH by using fluorescence-based DNA sequencing technology. RESULTS: Of 16 RB cases, 13 showed LOH on chromosome 13. The frequency of LOH on 13q14 was about 75% (12/16), consistent with other reports. Investigation of parental origin of lost RB1 alleles showed that, in all these cases, the paternal alleles were preferentially lost. Aside from the RB1 locus, other regions with the frequency of LOH above 30% in these tumors were D13S265, D13S158, D13S170, D13S218, D13S285 and D13S159. In particular, the D13S265 locus at 13q31-32 showed the highest rate of allele loss (64%, 9/14 informative cases), suggesting the presence of 1 or several genes whose loss of function may contribute to the development of RB. Comparison of the genotypic characteristics of 3 sites of frequent LOH (D13S153, D13S263 and D13S265) with the clinicopathological phenotype, respectively, showed that LOH of each locus was preferentially associated with a significantly younger age at diagnosis of RB. CONCLUSIONS: LOH analysis at some specific loci on chromosome 13 may be of a value in RB patients as diagnostic markers.