Cloning and characterization of an autonomous replication sequence from Coxiella burnetii.
Identifieur interne : 004296 ( Main/Exploration ); précédent : 004295; suivant : 004297Cloning and characterization of an autonomous replication sequence from Coxiella burnetii.
Auteurs : M. Suhan ; S Y Chen ; H A Thompson ; T A Hoover ; A. Hill ; J C WilliamsSource :
- Journal of bacteriology [ 0021-9193 ] ; 1994.
Descripteurs français
- KwdFr :
- ADN bactérien (génétique), ADN bactérien (isolement et purification), Amorces ADN, Cartographie de restriction, Clonage moléculaire (), Coxiella burnetii (génétique), Données de séquences moléculaires, Escherichia coli, Facteurs R (génétique), Plasmides, Réaction de polymérisation en chaîne, Réplication de l'ADN (génétique), Résistance à la kanamycine (génétique), Séquence nucléotidique, Vecteurs génétiques, Électrophorèse bidimensionnelle sur gel, Électrophorèse sur gel d'agar.
- MESH :
- génétique : ADN bactérien, Coxiella burnetii, Facteurs R, Réplication de l'ADN, Résistance à la kanamycine.
- isolement et purification : ADN bactérien.
- Amorces ADN, Cartographie de restriction, Clonage moléculaire, Données de séquences moléculaires, Escherichia coli, Plasmides, Réaction de polymérisation en chaîne, Séquence nucléotidique, Vecteurs génétiques, Électrophorèse bidimensionnelle sur gel, Électrophorèse sur gel d'agar.
English descriptors
- KwdEn :
- Base Sequence, Cloning, Molecular (methods), Coxiella burnetii (genetics), DNA Primers, DNA Replication (genetics), DNA, Bacterial (genetics), DNA, Bacterial (isolation & purification), Electrophoresis, Agar Gel, Electrophoresis, Gel, Two-Dimensional, Escherichia coli, Genetic Vectors, Kanamycin Resistance (genetics), Molecular Sequence Data, Plasmids, Polymerase Chain Reaction, R Factors (genetics), Restriction Mapping.
- MESH :
- chemical , genetics : DNA, Bacterial.
- chemical , isolation & purification : DNA, Bacterial.
- chemical : DNA Primers.
- genetics : Coxiella burnetii, DNA Replication, Kanamycin Resistance, R Factors.
- methods : Cloning, Molecular.
- Base Sequence, Electrophoresis, Agar Gel, Electrophoresis, Gel, Two-Dimensional, Escherichia coli, Genetic Vectors, Molecular Sequence Data, Plasmids, Polymerase Chain Reaction, Restriction Mapping.
Abstract
A Coxiella burnetii chromosomal fragment capable of functioning as an origin for the replication of a kanamycin resistance (Kanr) plasmid was isolated by use of origin search methods utilizing an Escherichia coli host. The 5.8-kb fragment was subcloned into phagemid vectors and was deleted progressively by an exonuclease III-S1 technique. Plasmids containing progressively shorter DNA fragments were then tested for their capability to support replication by transformation of an E. coli polA strain. A minimal autonomous replication sequence (ARS) was delimited to 403 bp. Sequencing of the entire 5.8-kb region revealed that the minimal ARS contained two consensus DnaA boxes, three A + T-rich 21-mers, a transcriptional promoter leading rightwards, and potential integration host factor and factor of inversion stimulation binding sites. Database comparisons of deduced amino acid sequences revealed that open reading frames located around the ARS were homologous to genes often, but not always, found near bacterial chromosomal origins; these included identities with rpmH and rnpA in E. coli and identities with the 9K protein and 60K membrane protein in E. coli and Pseudomonas species. These and direct hybridization data suggested that the ARS was chromosomal and not associated with the resident plasmid QpH1. Two-dimensional agarose gel electrophoresis did not reveal the presence of initiating intermediates, indicating that the ARS did not initiate chromosome replication during laboratory growth of C. burnetii.
DOI: 10.1128/jb.176.17.5233-5243.1994
PubMed: 8071197
Affiliations:
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Suhan</name><affiliation><nlm:affiliation>Department of Microbiology and Immunology, West Virginia University, Morgantown 26506-9177.</nlm:affiliation><wicri:noCountry code="subField">Morgantown 26506-9177</wicri:noCountry></affiliation></author><author><name sortKey="Chen, S Y" sort="Chen, S Y" uniqKey="Chen S" first="S Y" last="Chen">S Y Chen</name></author><author><name sortKey="Thompson, H A" sort="Thompson, H A" uniqKey="Thompson H" first="H A" last="Thompson">H A Thompson</name></author><author><name sortKey="Hoover, T A" sort="Hoover, T A" uniqKey="Hoover T" first="T A" last="Hoover">T A Hoover</name></author><author><name sortKey="Hill, A" sort="Hill, A" uniqKey="Hill A" first="A" last="Hill">A. Hill</name></author><author><name sortKey="Williams, J C" sort="Williams, J C" uniqKey="Williams J" first="J C" last="Williams">J C Williams</name></author></analytic><series><title level="j">Journal of bacteriology</title><idno type="ISSN">0021-9193</idno><imprint><date when="1994" type="published">1994</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Base Sequence</term><term>Cloning, Molecular (methods)</term><term>Coxiella burnetii (genetics)</term><term>DNA Primers</term><term>DNA Replication (genetics)</term><term>DNA, Bacterial (genetics)</term><term>DNA, Bacterial (isolation & purification)</term><term>Electrophoresis, Agar Gel</term><term>Electrophoresis, Gel, Two-Dimensional</term><term>Escherichia coli</term><term>Genetic Vectors</term><term>Kanamycin Resistance (genetics)</term><term>Molecular Sequence Data</term><term>Plasmids</term><term>Polymerase Chain Reaction</term><term>R Factors (genetics)</term><term>Restriction Mapping</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN bactérien (génétique)</term><term>ADN bactérien (isolement et purification)</term><term>Amorces ADN</term><term>Cartographie de restriction</term><term>Clonage moléculaire ()</term><term>Coxiella burnetii (génétique)</term><term>Données de séquences moléculaires</term><term>Escherichia coli</term><term>Facteurs R (génétique)</term><term>Plasmides</term><term>Réaction de polymérisation en chaîne</term><term>Réplication de l'ADN (génétique)</term><term>Résistance à la kanamycine (génétique)</term><term>Séquence nucléotidique</term><term>Vecteurs génétiques</term><term>Électrophorèse bidimensionnelle sur gel</term><term>Électrophorèse sur gel d'agar</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" type="chemical" xml:lang="en"><term>DNA Primers</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Coxiella burnetii</term><term>DNA Replication</term><term>Kanamycin Resistance</term><term>R Factors</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ADN bactérien</term><term>Coxiella burnetii</term><term>Facteurs R</term><term>Réplication de l'ADN</term><term>Résistance à la kanamycine</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>ADN bactérien</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Cloning, Molecular</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Base Sequence</term><term>Electrophoresis, Agar Gel</term><term>Electrophoresis, Gel, Two-Dimensional</term><term>Escherichia coli</term><term>Genetic Vectors</term><term>Molecular Sequence Data</term><term>Plasmids</term><term>Polymerase Chain Reaction</term><term>Restriction Mapping</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Amorces ADN</term><term>Cartographie de restriction</term><term>Clonage moléculaire</term><term>Données de séquences moléculaires</term><term>Escherichia coli</term><term>Plasmides</term><term>Réaction de polymérisation en chaîne</term><term>Séquence nucléotidique</term><term>Vecteurs génétiques</term><term>Électrophorèse bidimensionnelle sur gel</term><term>Électrophorèse sur gel d'agar</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">A Coxiella burnetii chromosomal fragment capable of functioning as an origin for the replication of a kanamycin resistance (Kanr) plasmid was isolated by use of origin search methods utilizing an Escherichia coli host. The 5.8-kb fragment was subcloned into phagemid vectors and was deleted progressively by an exonuclease III-S1 technique. Plasmids containing progressively shorter DNA fragments were then tested for their capability to support replication by transformation of an E. coli polA strain. A minimal autonomous replication sequence (ARS) was delimited to 403 bp. Sequencing of the entire 5.8-kb region revealed that the minimal ARS contained two consensus DnaA boxes, three A + T-rich 21-mers, a transcriptional promoter leading rightwards, and potential integration host factor and factor of inversion stimulation binding sites. Database comparisons of deduced amino acid sequences revealed that open reading frames located around the ARS were homologous to genes often, but not always, found near bacterial chromosomal origins; these included identities with rpmH and rnpA in E. coli and identities with the 9K protein and 60K membrane protein in E. coli and Pseudomonas species. These and direct hybridization data suggested that the ARS was chromosomal and not associated with the resident plasmid QpH1. Two-dimensional agarose gel electrophoresis did not reveal the presence of initiating intermediates, indicating that the ARS did not initiate chromosome replication during laboratory growth of C. burnetii.</div></front></TEI><affiliations><list></list><tree><noCountry><name sortKey="Chen, S Y" sort="Chen, S Y" uniqKey="Chen S" first="S Y" last="Chen">S Y Chen</name><name sortKey="Hill, A" sort="Hill, A" uniqKey="Hill A" first="A" last="Hill">A. Hill</name><name sortKey="Hoover, T A" sort="Hoover, T A" uniqKey="Hoover T" first="T A" last="Hoover">T A Hoover</name><name sortKey="Suhan, M" sort="Suhan, M" uniqKey="Suhan M" first="M" last="Suhan">M. Suhan</name><name sortKey="Thompson, H A" sort="Thompson, H A" uniqKey="Thompson H" first="H A" last="Thompson">H A Thompson</name><name sortKey="Williams, J C" sort="Williams, J C" uniqKey="Williams J" first="J C" last="Williams">J C Williams</name></noCountry></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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