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Related Concept Videos

Mechanism of Conjugation01:19

Mechanism of Conjugation

403
Bacterial conjugation is a mechanism of horizontal gene transfer that enables the exchange of genetic material between bacterial cells through direct contact. This process is facilitated by a donor cell carrying a conjugative plasmid, which encodes genes necessary for pilus formation, DNA replication, and transfer. The conjugative plasmid plays a central role in initiating and executing the transfer of genetic material.The tra region of the conjugative plasmid encodes proteins responsible for...
403

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Automated Rational Strain Construction Based on High-Throughput Conjugation.

Niklas Tenhaef1, Robert Stella1, Julia Frunzke1

  • 1Institute of Bio- and Geosciences - IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich 52425, Germany.

ACS Synthetic Biology
|February 17, 2021
PubMed
Summary
This summary is machine-generated.

Automating molecular cloning for synthetic biology streamlines DNA assembly and expression. This study presents an automated workflow for rational plasmid construction and transfer into Corynebacterium glutamicum, enhancing biosensor development.

Keywords:
DNA assemblyhigh-throughput conjugationlaboratory automationmolecular cloningstrain constructiontranscription factor-based biosensors

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Area of Science:

  • Synthetic biology
  • Molecular biology
  • Biotechnology

Background:

  • Molecular cloning is fundamental to synthetic biology but often manual and time-consuming.
  • Automating rational DNA construction and expression workflows is challenging, especially for less tractable hosts.
  • Few automated workflows exist for rational design and expression in non-model organisms.

Purpose of the Study:

  • To develop an automated workflow for rational plasmid construction and conjugative transfer.
  • To integrate DNA assembly, expression, and host transfer into a single automated process.
  • To apply this workflow for creating and characterizing transcription factor-biosensors in Corynebacterium glutamicum.

Main Methods:

  • Development of an automated workflow integrating DNA synthesis, cloning, and transformation.
  • Utilized a custom-designed software tool to manage the automated process.
  • Constructed and screened a library of rationally designed Lrp transcription factor-biosensors.
  • Performed conjugative transfer into Corynebacterium glutamicum.

Main Results:

  • Successfully automated the rational construction and transfer of plasmids into Corynebacterium glutamicum.
  • Developed a library of Lrp-based transcription factor-biosensors.
  • Identified a biosensor with an improved dynamic range.
  • Provided evidence for a dual regulatory role of Lrp in Corynebacterium glutamicum.

Conclusions:

  • The automated workflow significantly accelerates rational plasmid construction and expression in synthetic biology.
  • This approach enables efficient development of functional biosensors in biotechnologically relevant organisms.
  • The study offers new insights into the regulatory mechanisms of Corynebacterium glutamicum Lrp.