Comparison of solution-based exome capture methods for next generation sequencing.
Comparison of solution-based exome capture methods for next generation sequencing.
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DOI：
10.1186/gb-2011-12-9-r94
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发表时间：
2011-09-28
期刊：
影响因子：
12.3
通讯作者：
Saarela J
中科院分区：
文献类型：
--
作者：
Sulonen AM;Ellonen P;Almusa H;Lepistö M;Eldfors S;Hannula S;Miettinen T;Tyynismaa H;Salo P;Heckman C;Joensuu H;Raivio T;Suomalainen A;Saarela J
使得能够在下一代测序仪器上对人类基因组的蛋白质编码序列进行靶向重测序的技术受到极大关注。我们对Agilent和Roche NimbleGen提供的基于溶液的外显子组捕获试剂盒进行了系统比较。用所有四种捕获方法捕获对照DNA样品并制备用于Illumina GAII测序。在比较中还使用了用相同方案制备的其他样品的序列数据。我们开发了一个生物信息学管道，用于序列数据的质量控制、短读段比对、变体鉴定和注释。在我们的分析中，来自NimbleGen捕获的高质量读取的百分比大于来自Agilent捕获的高质量读取的百分比，其与捕获靶区域对齐。在所有外显子组富集方法中，靶序列的高GC含量与捕获成功率差相关。杂合变体的平均等位基因平衡的比较表明，在所有方法中，在靶区域内的杂合变体位置中具有比变体碱基更多的参考碱基的趋势。与来自SNP阵列的基因型相比，基因型一致性几乎没有差异。当与全基因组关联阵列上的常见SNP相比时，需要至少11×覆盖度才能以99%的准确度进行杂合子基因型调用。用NimbleGen试剂盒捕获的文库与靶区域更准确地对齐。更新的NimbleGen试剂盒最有效地覆盖了外显子组，最小覆盖率为20×，但没有一个试剂盒捕获了所有共有编码序列注释的外显子。
Techniques enabling targeted re-sequencing of the protein coding sequences of the human genome on next generation sequencing instruments are of great interest. We conducted a systematic comparison of the solution-based exome capture kits provided by Agilent and Roche NimbleGen. A control DNA sample was captured with all four capture methods and prepared for Illumina GAII sequencing. Sequence data from additional samples prepared with the same protocols were also used in the comparison. We developed a bioinformatics pipeline for quality control, short read alignment, variant identification and annotation of the sequence data. In our analysis, a larger percentage of the high quality reads from the NimbleGen captures than from the Agilent captures aligned to the capture target regions. High GC content of the target sequence was associated with poor capture success in all exome enrichment methods. Comparison of mean allele balances for heterozygous variants indicated a tendency to have more reference bases than variant bases in the heterozygous variant positions within the target regions in all methods. There was virtually no difference in the genotype concordance compared to genotypes derived from SNP arrays. A minimum of 11× coverage was required to make a heterozygote genotype call with 99% accuracy when compared to common SNPs on genome-wide association arrays. Libraries captured with NimbleGen kits aligned more accurately to the target regions. The updated NimbleGen kit most efficiently covered the exome with a minimum coverage of 20×, yet none of the kits captured all the Consensus Coding Sequence annotated exons.
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DOI：
10.1126/science.1194472
发表时间：
2010-12-03
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Harbour JW;Onken MD;Roberson ED;Duan S;Cao L;Worley LA;Council ML;Matatall KA;Helms C;Bowcock AM
通讯作者：
Bowcock AM
影响因子：
3
作者：
Slater GS;Birney E
通讯作者：
Birney E
影响因子：
30.8
作者：
Hoischen, Alexander;van Bon, Bregje W. M.;Veltman, Joris A.
通讯作者：
Veltman, Joris A.
使用下一代 DNA 测序数据进行变异发现和基因分型的框架。
影响因子：
30.8
作者：
通讯作者：
--
影响因子：
12.3
作者：
Fisher S;Barry A;Abreu J;Minie B;Nolan J;Delorey TM;Young G;Fennell TJ;Allen A;Ambrogio L;Berlin AM;Blumenstiel B;Cibulskis K;Friedrich D;Johnson R;Juhn F;Reilly B;Shammas R;Stalker J;Sykes SM;Thompson J;Walsh J;Zimmer A;Zwirko Z;Gabriel S;Nicol R;Nusbaum C
通讯作者：
Nusbaum C