Coupled amplification and sequencing of genomic DNA.
Coupled amplification and sequencing of genomic DNA.
复制标题
基因组 DNA 的耦合扩增和测序。
DOI：
10.1073/pnas.88.7.2815
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发表时间：
1991
影响因子：
11.1
通讯作者：
Kidd,KK
中科院分区：
文献类型：
--
作者：
Ruano,G;Kidd,KK
在PCR的指数期期间添加双脱氧核糖核苷酸应导致两个互补序列梯的合成。我们已经探索了这一假设，以开发基因组DNA的偶联扩增和测序。偶联扩增和测序是一种在扩增时对模板的两条链进行测序的双相方法。阶段I从基因组DNA样品中选择并扩增单个靶标。第II阶段使用来自第I阶段反应的等分试样与末端标记的引物和双脱氧核苷酸混合，完成测序以及靶的额外扩增。我们已经成功地应用偶联扩增和测序的300碱基对片段4酶上游HOX2B直接从人类全基因组DNA。
Addition of dideoxyribonucleotides during the exponential phase of the PCR should result in the synthesis of two complementary sequence ladders. We have explored this hypothesis to develop coupled amplification and sequencing of genomic DNA. Coupled amplification and sequencing is a biphasic method for sequencing both strands of template as they are amplified. Stage I selects and amplifies a single target from the genomic DNA sample. Stage II accomplishes the sequencing as well as additional amplification of the target using aliquots from the stage I reaction mixed with end-labeled primer and dideoxynucleotides. We have successfully applied coupled amplification and sequencing to a 300-base-pair fragment 4 kilobases upstream from HOX2B directly from human whole genomic DNA.