The present disclosure provides compositions and methods that employ the compositions for conducting pairwise sequencing and for generating concatemer template molecules for pairwise sequencing. The concatemers can be generated using a rolling circle amplification reaction which is conducted either on-support, or conducted in-solution and then distributed onto a support. The rolling circle amplification reaction generates concatemers containing tandem copies of a sequence of interest and at least one universal adaptor sequence. An increase in the number of tandem copies in a given concatemer increases the number of sites along the concatemer for hybridizing to multiple sequencing primers which serve as multiple initiation sites for polymerase-catalyzed sequencing reactions. When the sequencing reaction employs detectably labeled nucleotides and/or detectably labeled multivalent molecules (e.g., having nucleotide units), the signals emitted by the nucleotides or nucleotide units that participate in the parallel sequencing reactions along the concatemer yields an increased signal intensity for each concatemer.
本公开提供了使用这些组合物进行配对测序和生成用于配对测序的连接子模板分子的组合物和方法。可以使用滚圆扩增反应生成连接体,该反应可以在支架上进行,也可以在溶液中进行,然后分布到支架上。滚圆扩增反应生成的同源物含有感兴趣序列和至少一个通用适
配体序列的串联拷贝。特定连接体中串联拷贝数量的增加会增加连接体上与多个测序引物杂交的位点数量,而测序引物是聚合酶催化测序反应的多个起始位点。当测序反应使用可检测标记的核苷酸和/或可检测标记的多价分子(例如,具有核苷酸单元)时,沿连接体参与平行测序反应的核苷酸或核苷酸单元发出的信号会增加每个连接体的信号强度。