Retrofitting the BAC cloning vector pBeloBAC11 by the insertion of a mutant loxP site

ABSTRACT A Salmonella enterica serovar Typhimurium strain that harbored a plasmid carrying a TEM-1-type β-lactamase gene was isolated from the blood and cerebrospinal fluid of an infant with meningitis. This 3.2-kb plasmid was further characterized to be a nonconjugative pGEM series cloning vector containing a foreign insert. The strain was likely laboratory derived and contaminated the environment before it caused the infection.

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Engineering a Minimal Cloning Vector from a pUC18 Plasmid Backbone with an Extended Multiple Cloning Site

10.20944/preprints201807.0287.v2 ◽

2018 ◽

Author(s):

Jens Staal ◽

Wouter De Schamphelaire ◽

Rudi Beyaert

Keyword(s):

Synthetic Biology ◽

Molecular Biology ◽

Restriction Enzyme ◽

Essential Role ◽

Building Blocks ◽

Cloning Vector ◽

Multiple Cloning Site ◽

Unique Restriction ◽

Plasmid Backbone

Minimal plasmids play an essential role in many intermediate steps in molecular biology. They can for example be used to assemble building blocks in synthetic biology or be used as intermediate cloning plasmids that are ideal for PCR-based mutagenesis methods. A small backbone also opens up for additional unique restriction enzyme cloning sites. Here we describe the generation of pICOz, a 1185 bp fully functional high-copy cloning plasmid with an extended multiple cloning site (MCS). To our knowledge, this is the smallest high-copy cloning vector ever described.

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A successful use of a new shuttle cloning vector pA13 for the cloning of the bacteriocins BacSJ and acidocin 8912

Archives of Biological Sciences ◽

10.2298/abs1002231k ◽

2010 ◽

Vol 62 (2) ◽

pp. 231-243 ◽

Cited By ~ 1

Author(s):

Milan Kojic ◽

Jelena Lozo ◽

B. Jovcic ◽

Ivana Strahinic ◽

D. Fira ◽

...

Keyword(s):

In Silico Analysis ◽

Functional Expression ◽

Open Reading Frames ◽

Cloning Vector ◽

The Novel ◽

Genetic Manipulations ◽

Genes Encoding ◽

Editorial Decision ◽

Terminal Amino

The aim of this paper was to research the molecular cloning of genes encoding the novel bacteriocin BacSJ from Lactobacillus paracasei subsp. paracasei BGSJ2-8 by using a newly constructed shuttle cloning vector pA13. A new shuttle-cloning vector, pA13, was constructed and successfully introduced into Escherichia coli, Lactobacillus and Lactococcus strains, showing a high segregational and structural stability in all three hosts. The natural plasmid pSJ2-8 from L. paracasei subsp. paracasei BGSJ2-8 was cloned in the pA13 using BamHI, obtaining the construct pB5. Sequencing and in silico analysis of the pB5 revealed 15 open reading frames (ORF). Plasmid pSJ2-8 harbors the genes encoding the production of two bacteriocins, BacSJ and acidocin 8912. The combined N-terminal amino acid sequencing of BacSJ in combination with DNA sequencing of the bacSJ2-8 gene enabled the determination of the primary structure of a bacteriocin BacSJ. The production and functional expression of BacSJ in homologous and heterologous hosts suggest that bacSJ2-8 and bacSJ2-8i together with the genes encoding the ABC transporter and accessory protein are the minimal requirement for the production of BacSJ. Biochemical and genetic analyses showed that BacSJ belongs to the class II bacteriocins. The shuttle cloning vector pA13 could be used as a tool for genetic manipulations in lactobacilli and lactococci. withdrawn; due to a printing error. Link to the Editorial Decision <a href="http://dx.doi.org/10.2298/ABS1004251U">10.2298/ABS1004251U</a>

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