Coupling single base extension to a spectral codification tool for increased throughput screening
详细信息    查看全文
文摘
We report a new strategy that combines a Förster Resonance Energy Transfer (FRET) based spectral codification tool with a single base extension (SBE) reaction for rapid and medium-throughput analysis of single nucleotide polymorphisms (SNPs). This strategy is based on the spectral codification – a donor (fluorophore labeled probe complementary to the region adjacent to an SNP) is used to induce specific FRET signatures from an acceptor fluorophore revealing the SNP variant. Using an SBE reaction and differently labeled ddNTPs, we can directly question each donor probe and retrieve information about which allele variant is present at that locus. The potential of the method is demonstrated by application to simultaneous questioning of two loci in the same reaction tube. Following calibration with all possible combinations of FRET pairs, an evaluation algorithm was calibrated so as to optimize base calling and allow unequivocal allele scoring with more than 80 % confidence (for two simultaneous loci being questioned, one homo- and one heterozygous). In conclusion, this spectral codification approach may constitute a solution towards increasing throughput capability of single base extension based assays.

© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号

地址:北京市海淀区学院路29号 邮编:100083

电话:办公室:(+86 10)66554848;文献借阅、咨询服务、科技查新:66554700