筛选和确认 SNP SNaPshot 多通路系统是 SNP 筛选和验证的理想工具。该试剂盒提供单管单碱基延伸⁄终止试剂,可用于标记 DNA 片段。使用 GeneScan-120 LIZ 片段大小标准品在任何 Applied Biosystems 毛细管电泳仪上运行 DNA 片段,以指示标记片段的大小。然后使用 GeneMapper 软件分析数据并生成等位基因调用。SNaPshot 多通路检测试剂盒可通过单管单毛细管形式在单碱基延伸过程中对多达10种引物-模板组合进行多通路检测。
评估 DNA 甲基化 甲基化⁄表观遗传学的研究正在成为癌症研究的重要组成部分。在检测甲基化的典型测定试剂盒中,DNA 的亚硫酸氢盐处理可使非甲基化胞嘧啶脱氨基并将其转化为尿嘧啶,而甲基化胞嘧啶保持不反应。PCR 扩增的后续步骤会将尿嘧啶碱基转化为胸腺嘧啶。使用 SNaPshot 检测已处理和未处理样品的碱基差异,以了解您样品的甲基化状态。
How many SNPs can the SNaPshot Multiplex kit analyze?
The SNaPshot Multiplex Kis is designed to interrogate up to 10 single nucleotide polymorphisms (SNPs) at known locations on 1-10 templates in a single tube.
I am observing unexpected peaks in my SNaPshot data. Why?
The extra peaks could be due to incomplete removal of PCR primers, incomplete removal of dNTPs from the PCR reaction, or incomplete removal of the fluorescently labeled ddNTPS from the SNaPshot reaction. Residual PCR primers and dNTPs will both participate in the SNaPshot reaction. For the PCR purification, please use fresh SAP and Exo1 or use another method of PCR purification. To address incomplete removal of fluorescently labeled ddNTPS, use fresh CIP or SAP.
If my product is a single-base extension of my primer, shouldn't my products be primer length +1?
Due to the influence of the dye on the mobility shift of the DNA fragments, the reported sizes will differ by a few bases from the actual sizes. This is particularly true with the shorter fragments as the relative contribution of the dye is greater. It is also recommended that you have a tail on the primer with a 5-7 bp difference between them to further separate the samples so there is no overlap.
My SNaPshot Multiplex Kit control reaction does not match the sizes given in the protocol. Is this normal?
This is normal as the SNaPshot control shown in the manual is from a 3100 Genetic Analyzer and POP-4 polymer. Dye-labeled DNA fragments can yield different sizes when run with a different instrument, polymer type, capillary array length, or size standard. Although the control sizes may not match that in the protocol, once the size has been established, the precision, or reproducibility will be consistent for a given fragment.