Article

Quantitative analysis of wobble splicing indicates that it is not tissue specific.

Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Genomics (impact factor: 3.02). 12/2006; 88(6):855-64. DOI:10.1016/j.ygeno.2006.07.004 pp.855-64
Source: PubMed

ABSTRACT Alternative splicing is an important mechanism mediating the function of genes in multicellular organisms. Recently, we discovered a new splicing-junction wobble mechanism that generates subtle alterations in mRNA by randomly selecting tandem 5' and 3' splicing-junction sites. Here we developed a sensitive approach to identify such splicing-junction wobble isoforms using polymerase chain reaction amplification with fluorescence-labeled primers encompassing the wobble-splicing boundary and capillary electrophoresis. Using the ING4 wobble isoforms as an example, we demonstrated that capillary electrophoresis can precisely separate DNA fragments with a small difference in size (<3 nt) and can be used to quantify the expression ratio, which thus measures the distribution of each splicing-junction wobble isoform in tissues. Based on our analyses of several genes, the relative ratio of each wobble-splicing isoform tends to be constant among various tissues. The occasional observed tissue heterogeneity of wobble-splicing transcripts can be generated only by genomic single-nucleotide polymorphisms around the splicing junction.

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Keywords

3' splicing-junction sites
 
Alternative splicing
 
capillary electrophoresis
 
generates subtle alterations
 
genes
 
genomic single-nucleotide polymorphisms
 
ING4 wobble isoforms
 
multicellular organisms
 
new splicing-junction wobble mechanism
 
occasional
 
polymerase chain reaction amplification
 
small difference
 
splicing junction
 
splicing-junction wobble isoform
 
splicing-junction wobble isoforms
 
tandem 5'
 
various tissues
 
wobble-splicing boundary
 
wobble-splicing isoform
 
wobble-splicing transcripts
 

Kuo-Wang Tsai