Poly(T) Variation in Heteroderid Nematode Mitochondrial Genomes is Predominantly an Artefact of Amplification
Poly(T) Variation in Heteroderid Nematode Mitochondrial Genomes is Predominantly an Artefact of Amplification
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DOI：
10.1007/s00239-010-9414-3
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发表时间：
2011-02-01
影响因子：
3.9
通讯作者：
Dowton, Mark
中科院分区：
文献类型：
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作者：
Riepsamen, Angelique H.;Gibson, Tracey;Dowton, Mark
我们评估了在体外聚合酶的错误率在多聚胸苷[聚（T）]束的线粒体DNA（mtDNA）的异皮线虫（Heterodera cajani）。这些线虫的线粒体DNA含有异常高数量的聚（T）区，以前曾被认为含有生物聚（T）长度变异。然而，使用克隆的分子，我们观察到，聚（T）的变化是在体外含有超过六个连续的T的区域。该人为错误率估计为7.3 × 10（-5）indels/poly（T）tract > 6 Ts/循环。然后将该速率与扩增生物样品后检测到的聚（T）变异速率进行比较，以估计聚（T）变异的“生物+人为”速率。人工和人工+生物率之间没有显著差异，表明生物样本中的大多数聚（T）变异是人工的。然后，我们研究了产生的聚（T）的变化，在一系列的模板与道长达16 Ts，利用一系列的Heteroderidae物种。我们观察到T缺失发生的频率比插入高五倍，并且随着多聚（T）区长度的增加，错误率有增加的趋势。这些发现对涉及具有许多均聚物片段的基因组的研究具有重要意义。
We assessed the rate of in vitro polymerase errors at polythymidine [poly(T)] tracts in the mitochondrial DNA (mtDNA) of a heteroderid nematode (Heterodera cajani). The mtDNA of these nematodes contain unusually high numbers of poly(T) tracts, and have previously been suggested to contain biological poly(T) length variation. However, using a cloned molecule, we observed that poly(T) variation was generated in vitro at regions containing more than six consecutive Ts. This artefactual error rate was estimated at 7.3 x 10(-5) indels/poly(T) tract > 6 Ts/cycle. This rate was then compared to the rate of poly(T) variation detected after the amplification of a biological sample, in order to estimate the 'biological + artefactual' rate of poly(T) variation. There was no significant difference between the artefactual and the artefactual + biological rates, suggesting that the majority of poly(T) variation in the biological sample was artefactual. We then examined the generation of poly(T) variation in a range of templates with tracts up to 16 Ts long, utilizing a range of Heteroderidae species. We observed that T deletions occurred five times more frequently than insertions, and a trend towards increasing error rates with increasing poly(T) tract length. These findings have significant implications for studies involving genomes with many homopolymer tracts.