Circumventing heterozygosity: sequencing the amplified genome of a single haploid Drosophila melanogaster embryo.
Circumventing heterozygosity: sequencing the amplified genome of a single haploid Drosophila melanogaster embryo.
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DOI：
10.1534/genetics.111.127530
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发表时间：
2011-06
期刊：
影响因子：
3.3
通讯作者：
Stevens K
中科院分区：
文献类型：
--
作者：
Langley CH;Crepeau M;Cardeno C;Corbett-Detig R;Stevens K
杂合性是高效、高质量的基因组组装和全基因组多态性和趋异性调查的主要挑战。在果蝇中，来自赤道种群的品系对近亲繁殖特别有抗性，因此对该模式生物的自然种群中的基因组变异的测定和分析施加了主要障碍。在这里，我们提出了一个简单的基因组测序方案的基础上全基因组扩增的雌核发育衍生的单倍体基因组的后代的女性交配的男性纯合子隐性雄性不育突变，ms（3）K81。配对末端序列的单个“泳道”（2 × 76 bp）提供了具有>95%高质量覆盖率（超过5个读段）的良好同线组装。基因组DNA的扩增适度扩大了基因组常染色质部分覆盖率的变化。它还增加了嵌合克隆的频率。但是嵌合克隆的低频率和随机基因组分布限制了它们对最终组装的影响。该方法为群体基因组测序提供了一条坚实的道路，并为许多其他系统提供了应用，其中少量基因组DNA具有独特的实验相关性。
Heterozygosity is a major challenge to efficient, high-quality genomic assembly and to the full genomic survey of polymorphism and divergence. In Drosophila melanogaster lines derived from equatorial populations are particularly resistant to inbreeding, thus imposing a major barrier to the determination and analyses of genomic variation in natural populations of this model organism. Here we present a simple genome sequencing protocol based on the whole-genome amplification of the gynogenetically derived haploid genome of a progeny of females mated to males homozygous for the recessive male sterile mutation, ms(3)K81. A single “lane” of paired-end sequences (2 × 76 bp) provides a good syntenic assembly with >95% high-quality coverage (more than five reads). The amplification of the genomic DNA moderately inflates the variation in coverage across the euchromatic portion of the genome. It also increases the frequency of chimeric clones. But the low frequency and random genomic distribution of the chimeric clones limits their impact on the final assemblies. This method provides a solid path forward for population genomic sequencing and offers applications to many other systems in which small amounts of genomic DNA have unique experimental relevance.
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