DOI 10.1186/s12934-015-0325-2

Precise cloning and tandem integration of large polyketide biosynthetic gene cluster using Streptomyces artificial chromosome system Hee‑Ju Nah, Min‑Woo Woo, Si‑Sun Choi and Eung‑Soo Kim* Background: Direct cloning combined with heterologous expression of a secondary metabolite biosynthetic gene cluster has become a useful strategy for production improvement and pathway modification of potentially valuable natural products present at minute quantities in original isolates of actinomycetes. However, precise cloning and effi‑ cient overexpression of an entire biosynthetic gene cluster remains challenging due to the ineffectiveness of current genetic systems in manipulating large‑sized gene clusters for heterologous as well as homologous expression. Results: A versatile Escherichia coli‑Streptomyces shuttle bacterial artificial chromosomal (BAC) conjugation vector, pSBAC, was used along with a cluster tandem integration approach to carry out homologous and heterologous over‑ expression of a large 80‑kb polyketide biosynthetic pathway gene cluster of tautomycetin (TMC), which is a protein phosphatase PP1/PP2A inhibitor and T cell‑specific immunosuppressant. Unique XbaI restriction sites were precisel