scholarly journals Concerted Assembly and Cloning of Multiple DNA Segments Using In Vitro Site-Specific Recombination: Functional Analysis of Multi-Segment Expression Clones

2004 ◽  
Vol 14 (10b) ◽  
pp. 2111-2120 ◽  
Author(s):  
D. L. Cheo
Keyword(s):  
Functional Analysis ◽  
Site Specific ◽  
Site Specific Recombination

Site-Specific Recombination Promoted in Vitro by the FLP Protein of the Yeast Two-Micron Plasmid

1986 ◽  
pp. 397-405
Author(s):  
Julie F. Senecoff ◽  
Robert C. Bruckner ◽  
Leslie Meyer-Leon ◽  
Cynthia A. Gates ◽  
Elizabeth Wood ◽  
...  
Keyword(s):  
Site Specific ◽  
Site Specific Recombination

scholarly journals Characterization of the Chromosome Dimer Resolution Site in Caulobacter crescentus

2019 ◽  
Vol 201 (24) ◽  
Author(s):  
Ali Farrokhi ◽  
Hua Liu ◽  
George Szatmari
Keyword(s):  
Cell Division ◽  
Caulobacter Crescentus ◽  
Consensus Sequence ◽  
Markov Modeling ◽  
Control Of Gene Expression ◽  
Site Specific ◽  
Content Type ◽  
Site Specific Recombination ◽  
Resolution System

ABSTRACT Chromosome dimers occur in bacterial cells as a result of the recombinational repair of DNA. In most bacteria, chromosome dimers are resolved by XerCD site-specific recombination at the dif (deletion-induced filamentation) site located in the terminus region of the chromosome. Caulobacter crescentus, a Gram-negative oligotrophic bacterium, also possesses Xer recombinases, called CcXerC and CcXerD, which have been shown to interact with the Escherichia coli dif site in vitro. Previous studies on Caulobacter have suggested the presence of a dif site (referred to in this paper as dif1CC), but no in vitro data have shown any association with this site and the CcXer proteins. Using recursive hidden Markov modeling, another group has proposed a second dif site, which we call dif2CC, which shows more similarity to the dif consensus sequence. Here, by using a combination of in vitro experiments, we compare the affinities and the cleavage abilities of CcXerCD recombinases for both dif sites. Our results show that dif2CC displays a higher affinity for CcXerC and CcXerD and is bound cooperatively by these proteins, which is not the case for the original dif1CC site. Furthermore, dif2CC nicked substrates are more efficiently cleaved by CcXerCD, and deletion of the site results in about 5 to 10% of cells showing an altered cellular morphology. IMPORTANCE Bacteria utilize site-specific recombination for a variety of purposes, including the control of gene expression, acquisition of genetic elements, and the resolution of dimeric chromosomes. Failure to resolve dimeric chromosomes can lead to cell division defects in a percentage of the cell population. The work presented here shows the existence of a chromosomal resolution system in C. crescentus. Defects in this resolution system result in the formation of chains of cells. Further understanding of how these cells remain linked together will help in the understanding of how chromosome segregation and cell division are linked in C. crescentus.

scholarly journals Xer-mediated site-specific recombination in vitro.

1996 ◽  
Vol 15 (5) ◽  
pp. 1172-1181 ◽  
Author(s):  
S. D. Colloms ◽  
R. McCulloch ◽  
K. Grant ◽  
L. Neilson ◽  
D. J. Sherratt
Keyword(s):  
Site Specific ◽  
Site Specific Recombination

scholarly journals Gin-mediated site-specific recombination in bacteriophage Mu DNA: overproduction of the protein and inversion in vitro

1984 ◽  
Vol 3 (10) ◽  
pp. 2415-2421 ◽  
Author(s):  
Gabriele Mertens ◽  
Andrea Hoffmann ◽  
Helmut Blöcker ◽  
Ronald Frank ◽  
Regine Kahmann
Keyword(s):  
Bacteriophage Mu ◽  
Site Specific ◽  
Site Specific Recombination ◽  
And Inversion

scholarly journals Functional Analysis of the Acinetobacter baumannii XerC and XerD Site-Specific Recombinases: Potential Role in Dissemination of Resistance Genes

Antibiotics ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 405
Author(s):  
David L. Lin ◽  
German M. Traglia ◽  
Rachel Baker ◽  
David J. Sherratt ◽  
Maria Soledad Ramirez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Functional Analysis ◽  
Acinetobacter Baumannii ◽  
Potential Role ◽  
Covalent Attachment ◽  
Recombination Reaction ◽  
Site Specific ◽  
Site Specific Recombination ◽  
E Coli ◽  
Resistant Genes

Modules composed of a resistance gene flanked by Xer site-specific recombination sites, the vast majority of which were found in Acinetobacter baumannii, are thought to behave as elements that facilitate horizontal dissemination. The A. baumannii xerC and xerD genes were cloned, and the recombinant clones used to complement the cognate Escherichia coli mutants. The complemented strains supported the resolution of plasmid dimers, and, as is the case with E. coli and Klebsiella pneumoniae plasmids, the activity was enhanced when the cells were grown in a low osmolarity growth medium. Binding experiments showed that the partially purified A. baumannii XerC and XerD proteins (XerCAb and XerDAb) bound synthetic Xer site-specific recombination sites, some of them with a nucleotide sequence deduced from existing A. baumannii plasmids. Incubation with suicide substrates resulted in the covalent attachment of DNA to a recombinase, probably XerCAb, indicating that the first step in the recombination reaction took place. The results described show that XerCAb and XerDAb are functional proteins and support the hypothesis that they participate in horizontal dissemination of resistant genes among bacteria.

scholarly journals Site-specific recombination of yeast 2-micron DNA in vitro.

1983 ◽  
Vol 80 (23) ◽  
pp. 7284-7288 ◽  
Author(s):  
D. Vetter ◽  
B. J. Andrews ◽  
L. Roberts-Beatty ◽  
P. D. Sadowski
Keyword(s):  
Site Specific ◽  
Site Specific Recombination

scholarly journals The Escherichia coli Fis Protein Stimulates Bacteriophage λ Integrative Recombination In Vitro

2003 ◽  
Vol 185 (10) ◽  
pp. 3076-3080 ◽  
Author(s):  
Dominic Esposito ◽  
Gary F. Gerard
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Essential Role ◽  
Bacteriophage Λ ◽  
Site Specific ◽  
Site Specific Recombination ◽  
E Coli ◽  
In Vivo Recombination

ABSTRACT The Escherichia coli nucleoid-associated protein Fis was previously shown to be involved in bacteriophage lambda site-specific recombination in vivo, enhancing the levels of both integrative recombination and excisive recombination. While purified Fis protein was shown to stimulate in vitro excision, Fis appeared to have no effect on in vitro integration reactions even though a 15-fold drop in lysogenization frequency had previously been observed in fis mutants. We demonstrate here that E. coli Fis protein does stimulate integrative lambda recombination in vitro but only under specific conditions which likely mimic natural in vivo recombination more closely than the standard conditions used in vitro. In the presence of suboptimal concentrations of Int protein, Fis stimulates the rate of integrative recombination significantly. In addition, Fis enhances the recombination of substrates with nonstandard topologies which may be more relevant to the process of in vivo phage lambda recombination. These data support the hypothesis that Fis may play an essential role in lambda recombination in the host cell.

scholarly journals Highly Efficient In Vitro Site-Specific Recombination System Based on Streptomyces Phage φBT1 Integrase

2008 ◽  
Vol 190 (19) ◽  
pp. 6392-6397 ◽  
Author(s):  
Lin Zhang ◽  
Xijun Ou ◽  
Guoping Zhao ◽  
Xiaoming Ding
Keyword(s):  
Dna Supercoiling ◽  
Site Specific ◽  
Significant Frequency ◽  
Site Specific Recombination ◽  
Recombination System ◽  
Efficient Integration ◽  
Rapid Kinetics ◽  
A Site ◽  
Site Specific Recombination System

ABSTRACT The Streptomyces phage φBT1 encodes a site-specific integrase of the large serine recombinase subfamily. In this report, the enzymatic activity of the φBT1 integrase was characterized in vitro. We showed that this integrase has efficient integration activity with substrate DNAs containing attB and attP sites, independent of DNA supercoiling or cofactors. Both intra- and intermolecular recombinations proceed with rapid kinetics. The recombination is highly specific, and no reactions are observed between pairs of sites including attB and attL, attB and attR, attP and attL, or attP and attR or between two identical att sequences; however, a low but significant frequency of excision recombination between attL and attR is observed in the presence of the φBT1 integrase alone. In addition, for efficient integration, the minimal sizes of attB and attP are 36 bp and 48 bp, respectively. This site-specific recombination system is efficient and simple to use; thus, it could have applications for the manipulation of DNA in vitro.

In Vitro Analysis of Hin-mediated Site-specific Recombination

1984 ◽  
Vol 49 (0) ◽  
pp. 751-760 ◽  
Author(s):  
R.C. Johnson ◽  
M.B. Bruist ◽  
M.B. Glaccum ◽  
M.I. Simon
Keyword(s):  
Site Specific ◽  
Site Specific Recombination ◽  
Vitro Analysis ◽  
In Vitro Analysis
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