Recent advances in the direct cloning of large natural product biosynthetic gene clusters

Large-scale genome-mining analyses have revealed that microbes potentially harbor a huge reservoir of uncharacterized natural product (NP) biosynthetic gene clusters (BGCs), and this has spurred a renaissance of novel drug discovery. However, the majority of these BGCs are often poorly or not at all expressed in their native hosts under laboratory conditions, and thus are regarded as silent/orphan BGCs. Currently, connecting silent BGCs to their corresponding NPs quickly and on a large scale is particularly challenging because of the lack of universal strategies and enabling technologies. Generally, the heterologous host-based genome mining strategy is believed to be a suitable alternative to the native host-based approach for prioritization of the vast and ever-increasing number of uncharacterized BGCs. In the last ten years, a variety of methods have been reported for the direct cloning of BGCs of interest, which is the first and rate-limiting step in the heterologous expression strategy. Essentially, each method requires that the following three issues be resolved: 1) how to prepare genomic DNA; 2) how to digest the bilateral boundaries for release of the target BGC; and 3) how to assemble the BGC and the capture vector. Here, we summarize recent reports regarding how to directly capture a BGC of interest and briefly discuss the advantages and disadvantages of each method, with an emphasis on the notion that direct cloning is very beneficial for accelerating genome mining research and large-scale drug discovery.