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Related Concept Videos

DNA-only Transposons02:57

DNA-only Transposons

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DNA-only transposons are called autonomous transposons since they code for the enzyme transposase that is required for the transposition mechanism. Insertion of transposons can alter gene functions in multiple ways. They can mutate the gene, alter gene expression by introducing a novel promoter or insulator sequence, introduce new splice sites, and change the mRNA transcripts produced, or remodel chromatin structure.
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Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
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Transposons make up a significant part of genomes of various organisms. Therefore, it is believed that transposition played a major evolutionary role in speciation by changing genome sizes and modifying gene expression patterns. For example, in bacteria, transposition can lead to conferring antibiotic resistance. Movement of transposable elements within the genetic pool of pathogenic bacteria can aid in transfer of antibiotic-resistant genetic elements. In eukaryotes, transposons can carry out...
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Transposons01:24

Transposons

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Transposons, or "jumping genes," are small mobile genetic elements (MGEs) that range from 700 to 40,000 base pairs in length. They are found in all organisms and can move within the same chromosome or transfer to different chromosomes. In some cases, transposons can also jump between different host DNA molecules, such as plasmids or viruses, contributing to genetic variability.Barbara McClintock first discovered these mobile genetic elements in the 1940s while studying maize genetics, and she...
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In-vitro Mutagenesis01:16

In-vitro Mutagenesis

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To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.
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Related Experiment Video

Updated: Mar 8, 2026

Generating Transposon Insertion Libraries in Gram-Negative Bacteria for High-Throughput Sequencing
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Improved genome sequencing using an engineered transposase.

Amirali Kia1, Christian Gloeckner2, Trina Osothprarop1

  • 1Department of Protein Engineering, Illumina Inc, 5200 Illumina Way, San Diego, CA, USA.

BMC Biotechnology
|January 19, 2017
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Summary

Protein engineering created a mutant transposase (Tn5-059) that reduces GC insertion bias. This innovation improves genome coverage uniformity and library diversity, leading to more efficient and cost-effective next-generation sequencing library preparation.

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Area of Science:

  • Genomics
  • Molecular Biology
  • Biotechnology

Background:

  • Next-generation sequencing (NGS) revolutionized genomic research with faster turnaround times and reduced costs.
  • Transposase-based methods, like Nextera, integrated DNA fragmentation and barcoding for rapid library preparation.
  • Existing Nextera methods exhibit a slight insertion bias, impacting uniformity.

Purpose of the Study:

  • To engineer a mutant transposase with reduced GC insertion bias.
  • To evaluate the performance of the engineered transposase in library preparation.
  • To assess improvements in genome coverage, library diversity, and DNA input tolerance.

Main Methods:

  • Protein engineering of a transposase enzyme.
  • Comparative analysis of Tn5-059 and Nextera v2 for library preparation.
  • Assessment of genome coverage uniformity and AT dropout rates.
  • Evaluation of library diversity and insert size consistency across varying DNA inputs.

Main Results:

  • Discovery of Tn5-059, a mutant transposase with significantly lowered GC insertion bias.
  • Tn5-059 demonstrated reduced AT dropout and enhanced genome coverage uniformity in bacterial and human genomes.
  • Higher library diversity and improved cost-efficiency per genome were observed for human exomes using Tn5-059.
  • Consistent library insert size was achieved with Tn5-059 across a tenfold range of DNA input quantities.

Conclusions:

  • The engineered Tn5-059 transposase offers improved uniformity and reduced AT dropout compared to existing methods.
  • Enhanced DNA input tolerance provides workflow flexibility and robustness.
  • Tn5-059 expands the utility of transposase-based library preparation for diverse applications like microbiome sequencing and chromatin profiling.