Comparison between Normalised and Unnormalised 454-Sequencing Libraries for Small-Scale RNA-Seq Studies.
Comparison between Normalised and Unnormalised 454-Sequencing Libraries for Small-Scale RNA-Seq Studies.
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DOI：
10.1155/2012/281693
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发表时间：
2012
影响因子：
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通讯作者：
Burke T
中科院分区：
文献类型：
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作者：
Ekblom R;Slate J;Horsburgh GJ;Birkhead T;Burke T
下一代转录组测序（RNA-Seq）在非模式生物的研究中应用越来越广泛。在这里，我们在对斑胸草雀的小规模研究中评估了在测序之前规范化cDNA文库的有效性。我们发现，与非规范化库相比，由规范化库生成的程序集具有更多的contigs，但使用更少的读取。使用规范化cDNA产生的contigs也可以检测到更多的基因，微卫星发现的效率高达73%。在正常化和非正常化cDNA中检测到的基因表达水平之间存在正相关，并且在正常化和非正常化文库中鉴定出的血液和脾脏之间差异表达的基因数量没有差异。我们得出的结论是，标准化cDNA文库更适合RNA-Seq的许多应用，这些文库也可用于定量基因表达研究。
Next-generation sequencing of transcriptomes (RNA-Seq) is being used increasingly in studies of nonmodel organisms. Here, we evaluate the effectiveness of normalising cDNA libraries prior to sequencing in a small-scale study of the zebra finch. We find that assemblies produced from normalised libraries had a larger number of contigs but used fewer reads compared to unnormalised libraries. Considerably more genes were also detected using the contigs produced from normalised cDNA, and microsatellite discovery was up to 73% more efficient in these. There was a positive correlation between the detected expression level of genes in normalised and unnormalised cDNA, and there was no difference in the number of genes identified as being differentially expressed between blood and spleen for the normalised and unnormalised libraries. We conclude that normalised cDNA libraries are preferable for many applications of RNA-Seq and that these can also be used in quantitative gene expression studies.