کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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98931 | 160913 | 2012 | 7 صفحه PDF | دانلود رایگان |
SE33 is one of the most informative markers in forensic use due to its high power of discrimination. During the course of developing the AmpFℓSTR® NGM SElect™ PCR Amplification Kit several SE33 primer designs were screened with one primer pair yielding a high frequency of discordant alleles when compared to the AmpFℓSTR® SEfiler Plus™ PCR Amplification Kit. This discordance was mostly specific to samples of African descent with an estimated frequency of 5.1% and was a result of a mobility shift of approximately +0.84 nt. The sequence analysis of the affected alleles revealed that the only difference from the wild type sequence was a single nucleotide polymorphism (SNP) outside of the SE33 repeat but within the amplicon of this particular set of experimental primers. In total, we identified three different SNPs all within 9 nt of each other, each of which could cause the mobility shift individually. Further characterization of this region via site directed mutagenesis and thermostability measurements strongly suggests that this polymorphic region contains a secondary structure that, when disrupted due to the presence of a variant SNP, results in a mobility shift relative to the wild type sequence. To overcome this problem, the SE33 primers used in the final configuration of the NGM SElect™ Kit avoided the amplification of this polymorphic region yielding in turn results highly concordant with the SEfiler Plus™ Kit.
Journal: Forensic Science International: Genetics - Volume 6, Issue 3, May 2012, Pages 310–316