High-throughput gene and SNP discovery in Eucalyptus grandis, an uncharacterized genome.
High-throughput gene and SNP discovery in Eucalyptus grandis, an uncharacterized genome.
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DOI：
10.1186/1471-2164-9-312
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发表时间：
2008-06-30
期刊：
影响因子：
4.4
通讯作者：
Kirst, Matias
中科院分区：
文献类型：
--
作者：
Novaes, Evandro;Drost, Derek R.;Farmerie, William G.;Pappas, Georgios J., Jr.;Grattapaglia, Dario;Sederoff, Ronald R.;Kirst, Matias
使用454焦磷酸测序技术的高通量测序的益处对于具有高社会或经济价值但基因组资源很少的物种可能最明显。利用这种新的测序技术进行基因测序和SNP发现的快速手段为基因组水平的研究提供了一套基线工具。然而，对于基本上没有先前基因序列信息的物种的大量短读段的测序将如何有效地支持重叠群组装和序列注释是值得怀疑的。为了对巨桉（Eucalyptus grandis）的基因序列进行首次广泛调查，我们利用454技术对148 Mbp的表达序列（EST）进行了测序和组装。EST序列产生从一个标准化的cDNA池组成的多个组织和基因型，促进发现同源的近一半的拟南芥基因，并在转录组的等位基因变异的全面调查。通过比对来自多个基因型的测序读数，我们检测到23，742个SNP，其中83%在样本中得到验证。使用修改的θ参数估计2，392个重叠群的全基因组核苷酸多样性，该参数适用于测量通过随机测序多基因型cDNA库检测到的多态性的遗传多样性。在非同义核苷酸的多样性估计平均4倍小于同义，这表明纯化选择。在2，001个重叠群中，非同义替换为同义替换（Ka/Ks）的平均值为0.30，并且向右倾斜，进一步支持大多数基因处于纯化选择下。这些估计的重叠群之间的比较确定了纯化和多样化选择下的基因的主要功能类别与以前的研究一致。在提供丰富的基础转录物序列，其中有限的先前基因组信息存在，这项工作创造了注释的E。美国能源部正在对grandis的基因组进行测序。此外，我们证明了用454焦磷酸测序大规模采样的SNP可用于检测基因之间的进化特征，为非模式植物物种的核苷酸多样性和Ka/Ks提供了第一个全基因组评估。
Benefits from high-throughput sequencing using 454 pyrosequencing technology may be most apparent for species with high societal or economic value but few genomic resources. Rapid means of gene sequence and SNP discovery using this novel sequencing technology provide a set of baseline tools for genome-level research. However, it is questionable how effective the sequencing of large numbers of short reads for species with essentially no prior gene sequence information will support contig assemblies and sequence annotation. With the purpose of generating the first broad survey of gene sequences in Eucalyptus grandis, the most widely planted hardwood tree species, we used 454 technology to sequence and assemble 148 Mbp of expressed sequences (EST). EST sequences were generated from a normalized cDNA pool comprised of multiple tissues and genotypes, promoting discovery of homologues to almost half of Arabidopsis genes, and a comprehensive survey of allelic variation in the transcriptome. By aligning the sequencing reads from multiple genotypes we detected 23,742 SNPs, 83% of which were validated in a sample. Genome-wide nucleotide diversity was estimated for 2,392 contigs using a modified theta (θ) parameter, adapted for measuring genetic diversity from polymorphisms detected by randomly sequencing a multi-genotype cDNA pool. Diversity estimates in non-synonymous nucleotides were on average 4x smaller than in synonymous, suggesting purifying selection. Non-synonymous to synonymous substitutions (Ka/Ks) among 2,001 contigs averaged 0.30 and was skewed to the right, further supporting that most genes are under purifying selection. Comparison of these estimates among contigs identified major functional classes of genes under purifying and diversifying selection in agreement with previous researches. In providing an abundance of foundational transcript sequences where limited prior genomic information existed, this work created part of the foundation for the annotation of the E. grandis genome that is being sequenced by the US Department of Energy. In addition we demonstrated that SNPs sampled in large-scale with 454 pyrosequencing can be used to detect evolutionary signatures among genes, providing one of the first genome-wide assessments of nucleotide diversity and Ka/Ks for a non-model plant species.
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影响因子：
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作者：
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影响因子：
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作者：
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作者：
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