کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1204155 | 1493640 | 2013 | 6 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: Ultrafast haplotyping of putative microRNA-binding sites in the WFS1 gene by multiplex polymerase chain reaction and capillary gel electrophoresis Ultrafast haplotyping of putative microRNA-binding sites in the WFS1 gene by multiplex polymerase chain reaction and capillary gel electrophoresis](/preview/png/1204155.png)
The transmembrane protein wolframin (WSF1) plays a crucial role in cell integrity in pancreatic beta cells and maintaining ER homeostasis. Genetic variations in the WFS1 gene have been described to be associated with Wolfram syndrome or type 2 diabetes mellitus. In this paper we report on an efficient double-tube allele-specific amplification method in conjunction with ultrafast capillary gel electrophoresis for direct haplotyping analysis of the SNPs in two important miRNA-binding sites (rs1046322 and rs9457) in the WFS1 gene. An automated single-channel capillary gel electrophoresis system was utilized in the method that provided dsDNA fragment analysis in less than 240 s. The light-emitting diode induced fluorescence (LEDIF) detection system enabled excellent sensitivity for automated haplotyping of a large number of clinical samples. The detection limit was 0.002 ng/μL using field amplified injection from water diluted samples. The dynamic quantitation range was 0.08–10.00 ng/μL (R2 = 0.9997) in buffer diluted samples.
► Simultaneous haplotyping of two miRNA-binding sites in the WSF1 is presented.
► A combination allele-specific amplification and capillary electrophoresis was used.
► Ultra-fast size determination of the generated PCR fragments was done by CGE.
► Excellent detection limit of 2 ng/ml was demonstrated.
Journal: Journal of Chromatography A - Volume 1286, 19 April 2013, Pages 229–234