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Full-Text Articles in Life Sciences
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
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 …
Pcr-Activated Cell Sorting As A General, Cultivation-Free Method For High-Throughput Identification And Enrichment Of Virus Hosts, Shaun W. Lim, Shea T. Lance, Kenneth M. Stedman, Adam R. Abate
Pcr-Activated Cell Sorting As A General, Cultivation-Free Method For High-Throughput Identification And Enrichment Of Virus Hosts, Shaun W. Lim, Shea T. Lance, Kenneth M. Stedman, Adam R. Abate
Biology Faculty Publications and Presentations
Characterizing virus-host relationships is critical for understanding the impact of a virus on an ecosystem, but is challenging with existing techniques, particularly for uncultivable species. We present a general, cultivation-free approach for identifying phage-associated bacterial cells. Using PCR-activated cell sorting, we interrogate millions of individual bacteria for the presence of specific phage nucleic acids. If the nucleic acids are present, the bacteria are recovered via sorting and their genomes analyzed. This allows targeted recovery of all possible host species in a diverse population associated with a specific phage, and can be easily targeted to identify the hosts of different phages …