Characterization of Holliday structures in FLP protein-promoted site-specific recombination

1990 ◽  
Vol 10 (1) ◽  
pp. 235-242
Author(s):  
L Meyer-Leon ◽  
R B Inman ◽  
M M Cox
Keyword(s):  
Reaction Rate ◽  
Reaction Pathway ◽  
Reaction Intermediates ◽  
Wild Type ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Isomerization Process ◽  
Previous Demonstration ◽  
Rate Limiting

Holliday structures are formed in the course of FLP protein-promoted site-specific recombination. Here, we demonstrate that Holliday structures are formed in reactions involving wild-type substrates and that they are kinetically competent with respect to the overall reaction rate. Together with a previous demonstration of chemical competence (L. Meyer-Leon, L.-C. Huang, S. W. Umlauf, M. M. Cox, and R. B. Inman, Mol. Cell. Biol. 8:3784-3796, 1988), Holliday structures therefore meet all criteria necessary to establish that they are obligate reaction intermediates in FLP-mediated site-specific recombination. In addition, kinetic evidence suggests that two distinct forms of the Holliday intermediate are present in the reaction pathway, interconverted in an isomerization process that is rate limiting at 0 degree C.

scholarly journals Characterization of Holliday structures in FLP protein-promoted site-specific recombination.

1990 ◽  
Vol 10 (1) ◽  
pp. 235-242 ◽  
Author(s):  
L Meyer-Leon ◽  
R B Inman ◽  
M M Cox
Keyword(s):  
Reaction Rate ◽  
Reaction Pathway ◽  
Reaction Intermediates ◽  
Wild Type ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Isomerization Process ◽  
Previous Demonstration ◽  
Rate Limiting

Holliday structures are formed in the course of FLP protein-promoted site-specific recombination. Here, we demonstrate that Holliday structures are formed in reactions involving wild-type substrates and that they are kinetically competent with respect to the overall reaction rate. Together with a previous demonstration of chemical competence (L. Meyer-Leon, L.-C. Huang, S. W. Umlauf, M. M. Cox, and R. B. Inman, Mol. Cell. Biol. 8:3784-3796, 1988), Holliday structures therefore meet all criteria necessary to establish that they are obligate reaction intermediates in FLP-mediated site-specific recombination. In addition, kinetic evidence suggests that two distinct forms of the Holliday intermediate are present in the reaction pathway, interconverted in an isomerization process that is rate limiting at 0 degree C.

scholarly journals Amplification of the Tetracycline Resistance Determinant of pAMα1 in Enterococcus faecalis Requires a Site-Specific Recombination Event Involving Relaxase

2002 ◽  
Vol 184 (18) ◽  
pp. 5187-5193 ◽  
Author(s):  
M. Victoria Francia ◽  
Don B. Clewell
Keyword(s):  
Enterococcus Faecalis ◽  
Recombination Event ◽  
Tandem Repeats ◽  
Tetracycline Resistance ◽  
Multicopy Plasmid ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Rate Limiting Step ◽  
A Site ◽  
Rate Limiting

ABSTRACT The small multicopy plasmid pAMα1 (9.75 kb) encoding tetracycline resistance in Enterococcus faecalis is known to generate tandem repeats of a 4.1-kb segment carrying tet(L) when cells are grown extensively in the presence of tetracycline. Here we show that the initial (rate-limiting) step involves a site-specific recombination event involving plasmid-encoded relaxase activity acting at two recombination sequences (RS1 and RS2) that flank the tet determinant. We also present the complete nucleotide sequence of pAMα1.

scholarly journals Identification of the Telomere elongation mutation in Drosophila

2018 ◽  
Author(s):  
Hemakumar M. Reddy ◽  
Thomas A. Randall ◽  
Radmila Capkova Frydrychova ◽  
James M. Mason
Keyword(s):  
Drosophila Melanogaster ◽  
Next Generation Sequencing ◽  
Ltr Retrotransposons ◽  
Wild Type ◽  
Dominant Mutation ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Telomere Elongation ◽  
Drosophila Genetic Reference Panel ◽  
Generation Sequencing

Background. Telomeres in Drosophila melanogaster are similar to those of other eukaryotes in terms of their function, although they are formed by non-LTR retrotransposons instead of telomerase-based short repeats. The length of the telomeres in Drosophila depends on the number of copies of these transposable elements. A dominant mutation, Tel1, causes a several-fold elongation of telomeres. Methods. In this study we identified the Tel1 mutation by a combination of transposon-induced, site-specific recombination and next generation sequencing. Results. Recombination located Tel1 to a 15 kb region in 92A. Comparison of the DNA sequence in this region with the Drosophila Genetic Reference Panel of wild type genomic sequences delimited Tel1 to a 3 bp deletion inside intron 8 of Ino80. Discussion. The mapped Tel1 mutation (3-bp deletion found in Ino80) did not appear to affect the quantity or length of the Ino80 transcript. Tel1 causes a significant reduction in transcripts of CG18493, a gene nested in an intron 8 of Ino80, which is expressed in ovaries and expected to encode a serine-type peptidase.

scholarly journals Advantages and disadvantages of conditional systems for characterization of essential genes in Toxoplasma gondii

Parasitology ◽  
2014 ◽  
Vol 141 (11) ◽  
pp. 1390-1398 ◽  
Author(s):  
ELENA JIMÉNEZ-RUIZ ◽  
ELEANOR H. WONG ◽  
GURMAN S. PALL ◽  
MARKUS MEISSNER
Keyword(s):  
Toxoplasma Gondii ◽  
High Throughput ◽  
Gene Function ◽  
Function Analysis ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Advantages And Disadvantages ◽  
Gene Function Analysis ◽  
Tetracycline Inducible System

SUMMARYThe dissection of apicomplexan biology has been highly influenced by the genetic tools available for manipulation of parasite DNA. Here, we describe different techniques available for the generation of conditional mutants. Comparison of the advantages and disadvantages of the three most commonly used regulation systems: the tetracycline inducible system, the regulation of protein stability and site-specific recombination are discussed. Using some previously described examples we explore some of the pitfalls involved in gene-function analysis using these systems that can lead to wrong or over-interpretation of phenotypes. We will also mention different options to standardize the application of these techniques for the characterization of gene function in high-throughput.

scholarly journals Characterization of pABVA01, a Plasmid Encoding the OXA-24 Carbapenemase from Italian Isolates of Acinetobacter baumannii

2009 ◽  
Vol 53 (8) ◽  
pp. 3528-3533 ◽  
Author(s):  
Marco Maria D'Andrea ◽  
Tommaso Giani ◽  
Silvia D'Arezzo ◽  
Alessandro Capone ◽  
Nicola Petrosillo ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
Acinetobacter Baumannii ◽  
Binding Sites ◽  
Inverted Repeats ◽  
Clonal Lineage ◽  
Recombination Mechanism ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Carbapenem Resistant

ABSTRACT Two epidemiologically unrelated carbapenem-resistant Acinetobacter baumannii isolates were investigated as representatives of the first Italian isolates producing the OXA-24 carbapenemase. Both isolates were of European clonal lineage II and carried an identical OXA-24-encoding plasmid, named pABVA01. Comparative analysis revealed that in pABVA01, bla OXA-24 was part of a DNA module flanked by conserved inverted repeats homologous to XerC/XerD binding sites, which in other Acinetobacter plasmids flank different DNA modules, suggesting mobilization by a novel site-specific recombination mechanism.

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