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[Construction and evaluation of a pUC-type prokaryotic promoter reporter system based on lacZ gene]. | LitMetric

[Construction and evaluation of a pUC-type prokaryotic promoter reporter system based on lacZ gene].

Sheng Wu Gong Cheng Xue Bao

Poultry Institute, Chinese Academy of Agricultural Sciences, Yangzhou 225125, Jiangsu, China.

Published: January 2021

AI Article Synopsis

  • Researchers developed a new prokaryotic promoter reporter system called pFGH, based on the lacZ gene, using a modified pUC replicon plasmid for broad applications in gene expression studies.
  • The pFGH06 plasmid exhibited significantly lower background activity than other variants and reference plasmids, making it ideal for testing both inducible and constitutive promoters in the MC4100 bacterial strain.
  • This system allows for efficient cloning and screening of promoter activities, boasting benefits like smaller size, multiple cloning sites, and adjustable background levels, enhancing its potential for use in various genetic research projects.

Article Abstract

To construct a prokaryotic promoter report system with wide applicability, a series of pFGH reporter vectors based on lacZ gene and pUC replicon were constructed from plasmid pFLX107 through the replacement of multiple cloning sites and sequence modifications. The plasmid with the lowest background activity was selected as the final report system with the lacZ gene deletion strain MC4100 as the host bacterium, following by testing with inducible promoter araBAD and the constitutive promoter rpsM. The background activity of pFGH06 was significantly lower than that of other plasmids of the same series, and even lower than that of reference plasmid pRCL at 28 °C (P<0.01). Further evaluation tests show that the plasmid pFGH06 could be used to clone and determine the activity of inducible promoter or constitutive promoter, and the complete recognition of the target promoter could be achieved through blue-white selection in the simulation test of promoter screening. Compared with the reported prokaryotic promoter report systems, pFGH06 has the advantages of smaller size, more multiple clone sites, adjustable background activity, high efficiency of promoter screening and recognition, thus with a wide application prospect.

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Source
http://dx.doi.org/10.13345/j.cjb.200262DOI Listing

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