BACKGROUND Next-generation sequencing (NGS) has transformed genomic research by reducing turnaround time and cost. However, no major breakthrough has been made in the upstream library preparation methods until the transposase-based Nextera method was invented. Nextera combines DNA fragmentation and barcoding in a single tube reaction and therefore enables a very fast workflow to sequencing-read...

Genes responsible for the utilization of benzoate, anthranilate or catechol (ben, ant, cat) of Pseudomonas aeruginosa PAO were mapped precisely using a cosmid clone carrying all these genes. Genes were localized either by subcloning and complementation or by Tn5 mutagenesis and mapping of the Tn5 insertions. To achieve this, a novel Tn5 mutagenesis procedure was developed by constructing a Tn5 ...

The ability to generate tagged mutants of Rhodococcus spp. will facilitate a deeper understanding of this medically and commercially important genus. The absence of efficient transposon systems in these organisms has here been overcome by the use of Tn5-based DNA-protein transposition complexes which can transpose at high efficiency. To achieve this, electroporation efficiencies and antibiotic ...

Coliphage P1 was used to transduce derivatives of transposons Tn5 and mini-Mu into marine Vibrio spp. Transposon Tn5 encoding tetracycline resistance (Tn5-132) was used to isolate mutants of Vibrio harveyi defective in genes for bioluminescence (lux). Insertion of transposon Tn5-132 into the lux gene region was demonstrated by intraspecific transduction with phage hv-1 and by Southern blot hybr...

The presence of transposons Tn10 or Tn5 in the genome increases the frequency with which a silent (promoter-less) gene (hisD) is mutationally activated. The activation frequency is increased 5-25-fold by Tn10 and 30-90-fold by Tn5. Activation of the hisD gene is achieved by transposition of the entire transposon or one of its flanking insertion sequences to a region just upstream of the silent ...

The TP120 plasmid is known to determine enhanced UV survival in E. coli wild type an uvrB and PolA mutants but not in RecA mutant. In order to analyze the function involved in the SOS repair, we have constructed a new plasmid named pR derived by cleavage of TP120 with Hind III endonuclease. This new plasmid maintains the Ap and UV resistance. The insertion of Tn5 transposon in the plasmid allow...

We have used a papillation screening technique to isolate mutations that increase the precise excision of insertion elements. The three mutations isolated stimulated precise excision of Tn5, Tn10, and the IS elements. They had a large, 20- to 600-fold, effect on excision of Tn5 at various chromosomal sites. The varied stimulation for different Tn5 insertions showed that the mutations altered th...

Prokaryotic transposon 5 (Tn5) serves as a model system for studying the molecular mechanism of DNA transposition. Elucidation of the X-ray co-crystal structure of Tn5 transposase complexed with a DNA recognition end sequence provided the first three-dimensional picture of an intermediate in a transposition/retroviral integration pathway. The many Tn5 transposase-DNA co-crystal structures now a...