title: A novel donor splice-site mutation of major intrinsic protein gene associated with congenital cataract in a Chinese family.
abstract: OBJECTIVE
  To identify the disease-causing gene in a Chinese family with autosomal dominant congenital cataract.
  
  
  METHODS
  Clinical and ophthalmologic examinations were performed on all members of a Chinese family with congenital cataract. Nine genes associated with congenital cataract were screened using direct DNA sequencing. Mutations were confirmed using restriction fragment length polymorphism (RFLP) analysis. The mutated multi-intronic plasmid (MIP) minigene, which carries the disease-causing splice-site mutation, and the wild-type (WT) MIP minigene were constructed using the pcDNA3.1 expression vector. Wild-type and mutant MIP minigene constructs were transiently transfected into HeLa cells. After 48 h of incubation at 37 °C, total RNA isolation and reverse transcription (RT)-PCR analysis were performed, and PCR products were separated and confirmed with sequencing.
  
  
  RESULTS
  Direct DNA sequence analysis identified a novel splice-site mutation in intron 3 (c.606+1 G>A) of the MIP gene. To investigate the manner in which the splice donor mutation could affect mRNA splicing, WT and mutant MIP minigenes were inserted in the pcDNA3.1 (+) vector. Constructs were transfected into HeLa cells. RT-PCR analysis showed that the donor splice site mutation led to deletion of exon 3 in the mRNA encoded by the MIP gene.
  
  
  CONCLUSIONS
  The present study identified a novel donor splice-site mutation (c.606+1G>A) in the MIP gene in a Chinese family with congenital cataract. In vitro RT-PCR analysis showed that this splice-site mutation resulted in the deletion of exon 3 from mRNA encoded by the MIP gene. This is the first report to show that donor splice-site mutation in MIP genes can cause autosomal dominant congenital cataract.
authors: Key Laboratory of Molecular Biophysics, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China.
authors: Lu
authors: L
authors: Zeng
authors: Wenqiang
authors: W
authors: Liu
authors: Wenguo
authors: W
authors: Feng
authors: Xing
authors: X
authors: Wang
authors: Hui
authors: H
authors: Dang
authors: Luna
authors: L
authors: Gao
authors: Jing
authors: J
authors: Yao
authors: Xianqin
authors: X
authors: Zhang
mesh_terms: D000328:Adult; D020346:Aquaporins / Q000235:genetics*; D044466:Asian People; D001483:Base Sequence; D002386:Cataract / Q000208:ethnology / Q000235:genetics* / Q000473:pathology; D002675:Child, Preschool; D005091:Exons; D005136:Eye Proteins / Q000235:genetics*; D005260:Female; D005799:Genes, Dominant; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D008297:Male; D008969:Molecular Sequence Data; D010375:Pedigree; D012150:Polymorphism, Restriction Fragment Length; D022821:RNA Splice Sites; D012326:RNA Splicing; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion
chemical_list: D020346:Aquaporins; D005136:Eye Proteins; D022821:RNA Splice Sites; C030328:aquaporin 0