Genetics and Genomics Commons^™

In Vivo Cloning Of Up To 16 Kb Plasmids In E. Coli Is As Simple As Pcr, Faqing Huang, Joseph Rankin Spengler, Allen Yang Huang

Faculty Publications

The precise assembly of defined DNA sequences into plasmids is an essential task in bioscience research. While a number of molecular cloning techniques have been developed, many methods require specialized expensive reagents or laborious experimental procedure. Not surprisingly, conventional cloning techniques based on restriction digestion and ligation are still commonly used in routine DNA cloning. Here, we describe a simple, fast, and economical cloning method based on RecA- and RecET-independent in vivo recombination of DNA fragments with overlapping ends using E. coli. All DNA fragments were prepared by a 2-consecutive PCR procedure with Q5 DNA polymerase and used …