Single-molecule sequencing resolves the detailed structure of complex satellite DNA loci in Drosophila melanogaster.
Single-molecule sequencing resolves the detailed structure of complex satellite DNA loci in Drosophila melanogaster.
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DOI：
10.1101/gr.213512.116
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发表时间：
2017-05
期刊：
影响因子：
7
通讯作者：
Larracuente AM
中科院分区：
文献类型：
--
作者：
Khost DE;Eickbush DG;Larracuente AM
高度重复的卫星DNA(SatDNA)重复序列存在于大多数真核生物基因组中。卫星DNA进化迅速，并在基因组稳定和染色体分离中发挥作用。它们的重复性给基因组组装带来了挑战，并使卫星DNA结构的详细研究变得困难。在这里，我们使用太平洋生物科学(PacBio)的单分子测序长读数(PacBio)来确定果蝇所有主要常染色体复合体SatDNA基因座的详细结构，特别是260-BP和Responder卫星。我们确定了产生这些先前未组装的卫星DNA基因座的高质量组装所需的最佳从头组装方法和参数组合，并使用分子和计算方法验证了这种组装。我们确定，计算密集型的PBcR-BLASR组装流水线比基于MHAP散列算法的更快、更高效的流水线产生更好的组装，验证重复基因座的组装是必不可少的。这些组装揭示了卫星DNA重复序列被组织成被转座元件打断的大阵列。序列中心的重复序列趋向于按顺序同质，这表明基因转换和不等交换通过协同进化导致重复均质，尽管不同复杂卫星座位的不等交换程度可能不同。我们在卫星DNA阵列中发现了高阶结构的证据，这表明最近的结构重排。这些组件为着丝粒周围异染色质中的卫星DNA的进化和功能基因组学提供了一个平台。
Highly repetitive satellite DNA (satDNA) repeats are found in most eukaryotic genomes. SatDNAs are rapidly evolving and have roles in genome stability and chromosome segregation. Their repetitive nature poses a challenge for genome assembly and makes progress on the detailed study of satDNA structure difficult. Here, we use single-molecule sequencing long reads from Pacific Biosciences (PacBio) to determine the detailed structure of all major autosomal complex satDNA loci in Drosophila melanogaster, with a particular focus on the 260-bp and Responder satellites. We determine the optimal de novo assembly methods and parameter combinations required to produce a high-quality assembly of these previously unassembled satDNA loci and validate this assembly using molecular and computational approaches. We determined that the computationally intensive PBcR-BLASR assembly pipeline yielded better assemblies than the faster and more efficient pipelines based on the MHAP hashing algorithm, and it is essential to validate assemblies of repetitive loci. The assemblies reveal that satDNA repeats are organized into large arrays interrupted by transposable elements. The repeats in the center of the array tend to be homogenized in sequence, suggesting that gene conversion and unequal crossovers lead to repeat homogenization through concerted evolution, although the degree of unequal crossing over may differ among complex satellite loci. We find evidence for higher-order structure within satDNA arrays that suggest recent structural rearrangements. These assemblies provide a platform for the evolutionary and functional genomics of satDNAs in pericentric heterochromatin.