Mariasegaram Maxy, Robinson Nicholas Andrew, Goddard Michael Edward
CSIRO Livestock Industries, Queensland Biosciences Precinct, 306 Carmody Road, St Lucia QLD 4067, Australia.
Genet Sel Evol. 2007 May-Jun;39(3):267-83. doi: 10.1186/1297-9686-39-3-267. Epub 2007 Apr 14.
This study represents the first attempt at an empirical evaluation of the DNA pooling methodology by comparing it to individual genotyping and interval mapping to detect QTL in a dairy half-sib design. The findings indicated that the use of peak heights from the pool electropherograms without correction for stutter (shadow) product and preferential amplification performed as well as corrected estimates of frequencies. However, errors were found to decrease the power of the experiment at every stage of the pooling and analysis. The main sources of errors include technical errors from DNA quantification, pool construction, inconsistent differential amplification, and from the prevalence of sire alleles in the dams. Additionally, interval mapping using individual genotyping gains information from phenotypic differences between individuals in the same pool and from neighbouring markers, which is lost in a DNA pooling design. These errors cause some differences between the markers detected as significant by pooling and those found significant by interval mapping based on individual selective genotyping. Therefore, it is recommended that pooled genotyping only be used as part of an initial screen with significant results to be confirmed by individual genotyping. Strategies for improving the efficiency of the DNA pooling design are also presented.
本研究首次尝试通过将DNA混合池方法与个体基因分型和区间作图进行比较，以在奶牛半同胞设计中检测数量性状基因座（QTL），从而对该方法进行实证评估。研究结果表明，在不校正拖尾（影子）产物和优先扩增的情况下，使用混合池电泳图中的峰高，其效果与校正后的频率估计值相当。然而，发现在混合池构建和分析的每个阶段，误差都会降低实验的效能。误差的主要来源包括DNA定量、混合池构建、不一致的差异扩增以及母畜中父本等位基因的流行等技术误差。此外，使用个体基因分型的区间作图可从同一混合池中个体之间的表型差异以及相邻标记中获取信息，而这些信息在DNA混合池设计中会丢失。这些误差导致通过混合池检测为显著的标记与基于个体选择性基因分型通过区间作图发现显著的标记之间存在一些差异。因此，建议将混合池基因分型仅用作初步筛选的一部分，显著结果需通过个体基因分型进行确认。本文还提出了提高DNA混合池设计效率的策略。
