• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Nucleus >Sequencing on the SOLiD 5500xl System – in-depth characterization of the GC bias
【24h】

Sequencing on the SOLiD 5500xl System – in-depth characterization of the GC bias

机译：在SOLiD 5500xl系统上进行测序– GC偏差的深入表征

获取原文
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Different types of sequencing biases have been described and subsequently improved for a variety of sequencing systems, mostly focusing on the widely used Illumina systems. Similar studies are missing for the SOLiD 5500xl system, a sequencer which produced many data sets available to researchers today. Describing and understanding the bias is important to accurately interpret and integrate these published data in various ongoing research projects. We report a particularly strong GC bias for this sequencing system when analyzing a defined gDNA mix of 5 microbes with a wide range of different GC contents (20–72%) when comparing to the expected distribution and Illumina MiSeq data from the same DNA pool. Since we observed this bias already under PCR-free conditions, changing the PCR conditions during library preparation – a common strategy to handle bias in the Illumina system - was not relevant. Source of the bias appeared to be an uneven heat distribution during the SOLiD emulsion PCR (ePCR) - for enrichment of libraries prior loading – since ePCR in either small pouches or in 96-well plates improved the GC bias.Sequencing of chromatin immunoprecipitated DNA (ChIP-seq) is a common approach in epigenetics. ChIP-seq of the mixed source histone mark H3K9ac (acetyl Histone H3 lysine 9), typically found on promoter regions and on gene bodies, including CpG islands, performed on a SOLiD 5500xl machine, resulted in major loss of reads at GC rich loci (GC content ≥ 62%), not explained by low sequencing depth. This was improved with adaptations of the ePCR.
机译：已经描述了不同类型的测序偏差，随后针对各种测序系统对其进行了改进，主要集中在广泛使用的Illumina系统上。 SOLiD 5500xl系统缺少类似的研究，SOLiD 5500xl系统产生了许多数据集，可供当今的研究人员使用。描述和理解偏差对于在各种正在进行的研究项目中准确地解释和整合这些已发布的数据非常重要。当与来自同一DNA池的预期分布和Illumina MiSeq数据进行比较时，当分析5种微生物的定义的gDNA混合物时，我们报告该测序系统的GC偏向特别强，该微生物具有多种不同的GC含量（20-72％）。由于我们已经在无PCR的条件下观察到这种偏倚，因此在文库制备过程中改变PCR条件（一种用于处理Illumina系统中偏倚的常见策略）并不重要。偏差的来源似乎是SOLiD乳液PCR（ePCR）期间的热量分布不均-在上样前富集文库-因为小袋或96孔板中的ePCR均可改善GC偏差。染色质免疫沉淀DNA测序（ ChIP-seq）是表观遗传学中的常用方法。混合来源组蛋白标记H3K9ac（乙酰基组蛋白H3赖氨酸9）的ChIP-seq通常在启动子区域和包括CpG岛的基因体上发现，在SOLiD 5500xl机器上进行，导致富含GC的基因座的读段大量丢失（ GC含量≥62％），低测序深度无法解释。通过ePCR的改进，这一点得到了改善。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号