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[Coding single nucleotide polymorphism is an ideal marker for detecting gene imprinting by 5' nuclease assay]
Mo-bin Wan1, Guan-shan Zhu, Rui-ying Zheng
1Department of Infectious Diseases, Changhai Hospital, Second Military Medical University, Shanghai, 200433 PR China. wanmb@online.sh.cn
Objective:
To establish a novel approach for quick and high throughput verification of human gene imprinting.
Methods:
By use of a pair of dye-labeled probes, 5' nuclease assay was combined with reverse transcriptase-PCR(RT-PCR) to genotype a coding single nucleotide polymorphism (cSNP), rs705(C/T) of a known imprinted gene, small nuclear ribonucleotide protein N (SNRPN), on both genomic DNA and cDNA of human lymphoblast cell lines.
Results:
Allele discrimination showed a clear monoallelic expression pattern of SNRPN, which was confirmed by RT-PCR based restriction fragment length polymorphisms. Pedigree analysis verified the paternal origin of expressed allele, which is in consistency with previous report.
Conclusion:
Coding SNP is an ideal marker for detecting gene imprinting by 5' nuclease assay. This approach has also a potentiality to discover differential allele expression of non-imprinted genes in order to find gene cis-acting functional polymorphism.