Fu D J, Broude N E, Köster H, Smith C L, Cantor C R
Sequenom, Inc., Boston, MA 02210, USA.
Proc Natl Acad Sci U S A. 1995 Oct 24;92(22):10162-6. doi: 10.1073/pnas.92.22.10162.
We have previously reported an enhanced version of sequencing by hybridization (SBH), termed positional SBH (PSBH). PSBH uses partially duplex probes containing single-stranded 3' overhangs, instead of simple single-stranded probes. Stacking interactions between the duplex probe and a single-stranded target allow us to reduce the probe sizes required to 5-base single-stranded overhangs. Here we demonstrate the use of PSBH to capture relatively long single-stranded DNA targets and perform standard solid-state Sanger sequencing on these primer-template complexes without ligation. Our results indicate that only 5 bases of known terminal sequence are required for priming. In addition, the partially duplex probes have the ability to capture their specific target from a mixture of five single-stranded targets with different 3'-terminal sequences. This indicates the potential utility of the PSBH approach to sequence mixtures of DNA targets without prior purification.
我们之前报道过一种杂交测序（SBH）的增强版本，称为定位SBH（PSBH）。PSBH使用含有单链3'突出端的部分双链探针，而不是简单的单链探针。双链探针与单链靶标之间的堆积相互作用使我们能够将所需的探针大小减小到5个碱基的单链突出端。在这里，我们展示了使用PSBH捕获相对较长的单链DNA靶标，并在这些引物-模板复合物上进行标准的固态桑格测序而无需连接。我们的结果表明，引发仅需要5个已知末端序列的碱基。此外，部分双链探针能够从具有不同3'末端序列的五个单链靶标的混合物中捕获其特定靶标。这表明PSBH方法在无需事先纯化的情况下对DNA靶标混合物进行测序的潜在实用性。
