Cloning of Circular DNAs from Microorganisms Using a Novel Plasmid Capture System

• Published in: Molecular biotechnology Vol. 44; no. 2; pp. 120 - 126
• Main Authors: Roh, Jong Yul, Wang, Yong, Liu, Qin, Tao, Xueying, Choi, Jae Young, Shim, Hee Jin, Xu, Hong Guang, Lee, Seungdon, Woo, Soo Dong, Jin, Byung Rae, Je, Yeon Ho
• Format: Journal Article
• Language: English
• Published: New York New York : Humana Press Inc 01.02.2010; Humana Press Inc; Springer; Springer Nature B.V
• Subjects: Autographa californica; Bacillus thuringiensis; Bacillus thuringiensis - genetics; Biochemistry; Biological and medical sciences; Biological Techniques; Biotechnology; Bracovirus; Cell Biology; Chemistry; Chemistry and Materials Science; Cloning; Cloning, Molecular - methods; Cotesia glomerata; Deoxyribonucleic acid; DNA; DNA, Circular - genetics; DNA, Viral - genetics; E coli; Escherichia coli; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Genomics; Human Genetics; Microorganisms; Models, Biological; Molecular biology; Mutagenesis; Nuclear polyhedrosis virus; Nucleopolyhedrovirus - genetics; Plasmids - genetics; Polymorphism; Protein Science; Circular DNA cloning; PCS-LZ; AcMNPV; In vitro transposition; Plasmid capture system (PCS); CgBV; Bacillus thuringiensis; Bacillaceae; In vitro; Plasmid; Transposition; Bacillales; DNA; Bacteria; Plasmid capture system (PCS) PCS-LZ; Microorganism
• ISSN: 1073-6085; 1559-0305
• DOI: 10.1007/s12033-009-9215-3

Plasmid capture system (PCS) facilitates cloning and manipulation of circular double-stranded DNA. We recently developed an improved PCS (PCS-LZ) to clone relatively large DNA molecules of 30-150 kb. The PCS-LZ donor consists of a mini-F replicon and a kanamycin resistance marker between Tn7 left and Tn7 right ends. Both the replicon and marker gene of the PCS-LZ donor are transferred into target plasmid DNAs by in vitro transposition, followed by replication in E. coli. Colonies are tested for lacZ expression by blue/white screening. Circular DNAs were obtained from plasmids of Bacillus thuringiensis, genome segments of Cotesia glomerata bracovirus and polymorphic genomes of Autographa californica nucleopolyhedrovirus. PCS-LZ is a powerful tool for use in genomic analysis and mutagenesis in microorganisms including invertebrate pathogens.