Simultaneous sequencing of multiple polymerase chain reaction products and combined polymerase chain reaction with cycle sequencing in single reactions.
Simultaneous sequencing of multiple polymerase chain reaction products and combined polymerase chain reaction with cycle sequencing in single reactions.
复制标题
多个聚合酶链反应产物的同时测序以及聚合酶链反应与单个反应中的循环测序的组合。
DOI：
10.1016/s0002-9440(10)64153-3
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发表时间：
2002
期刊：
影响因子：
--
通讯作者：
Eshleman,JamesR
中科院分区：
文献类型：
--
作者：
Murphy,KathleenM;Eshleman,JamesR
DNA sequencing is considered the gold standard for nucleic acid identification and mutation detection. However, sequencing is labor intensive because it requires previous amplification and only a single sequence is analyzed at a time. We developed two novel strategies that substantially improve DNA sequencing. The first allows multiple polymerase chain reaction (PCR) products to be sequenced in a single sequencing reaction and analyzed simultaneously in a single lane or capillary. Simultaneous sequencing by this method, designated “SimulSeq,” can provide either simultaneous single-direction sequencing of multiple genes or simultaneous forward and reverse sequencing from a single gene. In the second approach, designated “AmpliSeq,” we demonstrate a technique combining PCR amplification and sequencing in a single reaction that is analyzed in a single lane or capillary. We demonstrate combined PCR with short bidirectional sequencing, and combined PCR with unidirectional sequencing. We anticipate that these methods will have utility in research and clinical settings where panels of mutations or large numbers of samples are be analyzed and/or when turnaround time is critical.
