Medina Laura Smeldy Jurado, Muzzio Marina, Schwab Marisol, Costantino María Leticia Bravi, Barreto Guillermo, Bailliet Graciela
Laboratorio de Genética Molecular Poblacional, Instituto Multidisciplinario de Biología Celular (IMBICE), CCT- CONICET-La Plata/CICPBA, La Plata, Argentina.
Electrophoresis. 2014 Sep;35(17):2524-7. doi: 10.1002/elps.201400020. Epub 2014 Jul 14.
We designed an allele-specific amplification protocol to optimize Y-chromosome SNP typing, which is an unavoidable step for defining the phylogenetic status of paternal lineages. It allows the simultaneous highly specific definition of up to six mutations in a single reaction by amplification fragment length polymorphism (AFLP) without the need of specialized equipment, at a considerably lower cost than that based on single-base primer extension (SNaPshot™) technology or PCR-RFLP systems, requiring as little as 0.5 ng DNA and compatible with the small fragments characteristic of low-quality DNA. By designation of two primers recognizing the derived and ancestral state for each SNP, which can be differentiated by size by the addition of a noncomplementary nucleotide tail, we could define major Y clades E, F, K, R, Q, and subhaplogroups R1, R1a, R1b, R1b1b, R1b1c, J1, J2, G1, G2, I1, Q1a3, and Q1a3a1 through amplification fragments that ranged between 60 and 158bp.
我们设计了一种等位基因特异性扩增方案，以优化Y染色体单核苷酸多态性（SNP）分型，这是确定父系谱系系统发育地位时不可避免的一步。它能够通过扩增片段长度多态性（AFLP）在单个反应中同时对多达六个突变进行高度特异性的鉴定，无需专门设备，成本远低于基于单碱基引物延伸（SNaPshot™）技术或PCR-RFLP系统，所需DNA少至0.5 ng，并且与低质量DNA的小片段特征相兼容。通过为每个SNP指定两条识别衍生状态和祖先状态的引物，通过添加非互补核苷酸尾可按大小区分，我们能够通过60至158 bp的扩增片段确定主要的Y染色体分支E、F、K、R、Q以及亚单倍群R1、R1a、R1b、R1b1b、R1b1c、J1、J2、G1、G2、I1、Q1a3和Q1a3a1。
