Tatsumi Kaori, Nishimura Osamu, Itomi Kazu, Tanegashima Chiharu, Kuraku Shigehiro
Phyloinformatics Unit, RIKEN Center for Life Science Technologies, Kobe, Japan.
Biotechniques. 2015 May 1;58(5):253-7. doi: 10.2144/000114288. eCollection 2015 May.
In de novo genome sequencing, mate-pair reads are crucial for scaffolding assembled contigs. However, preparation of mate-pair libraries is not a trivial task, even when using one of the latest approaches, the Nextera Mate Pair Sample Prep Kit from Illumina. To reduce cost and enhance library yield and fidelity when using this kit, we have modified the manufacturer's protocol based on (i) variable tagmentation conditions, (ii) intensive DNA shearing to decrease library insert length, and (iii) sequencing on an Illumina HiSeq with >150 cycles. Finally, we provide additional suggestions for further improvement in the application of this kit.
在从头基因组测序中，配对末端读段对于拼接组装的重叠群至关重要。然而，即使使用最新方法之一（Illumina公司的Nextera Mate Pair Sample Prep Kit），制备配对末端文库也并非易事。为了在使用该试剂盒时降低成本、提高文库产量和保真度，我们基于以下几点修改了制造商的方案：（i）可变的转座酶切条件；（ii）高强度DNA剪切以缩短文库插入片段长度；（iii）在Illumina HiSeq上进行超过150个循环的测序。最后，我们为该试剂盒应用的进一步改进提供了其他建议。
