A genomic library-based amplification approach (GL-PCR) for the mapping of multiple IS6110 insertion sites and strain differentiation of Mycobacterium tuberculosis.
Identifieur interne : 000029 ( Main/Exploration ); précédent : 000028; suivant : 000030A genomic library-based amplification approach (GL-PCR) for the mapping of multiple IS6110 insertion sites and strain differentiation of Mycobacterium tuberculosis.
Auteurs : Amine Namouchi [Tunisie] ; Helmi MardassiSource :
- Journal of microbiological methods [ 0167-7012 ] ; 2006.
Descripteurs français
- KwdFr :
- ADN bactérien (composition chimique), ADN bactérien (génétique), Analyse de séquence d'ADN (MeSH), Banque génomique (MeSH), Cartographie chromosomique (méthodes), Clonage moléculaire (MeSH), Génome bactérien (génétique), Humains (MeSH), Mycobacterium tuberculosis (classification), Mycobacterium tuberculosis (génétique), Réaction de polymérisation en chaîne (MeSH), Transformation génétique (MeSH), Tuberculose (microbiologie), Type II site-specific deoxyribonuclease (métabolisme), Éléments transposables d'ADN (génétique).
- MESH :
- composition chimique : ADN bactérien, Mycobacterium tuberculosis.
- génétique : ADN bactérien, Génome bactérien, Mycobacterium tuberculosis, Éléments transposables d'ADN.
- microbiologie : Tuberculose.
- métabolisme : Type II site-specific deoxyribonuclease.
- méthodes : Cartographie chromosomique.
- Analyse de séquence d'ADN, Banque génomique, Clonage moléculaire, Humains, Réaction de polymérisation en chaîne, Transformation génétique.
English descriptors
- KwdEn :
- Chromosome Mapping (methods), Cloning, Molecular (MeSH), DNA Transposable Elements (genetics), DNA, Bacterial (chemistry), DNA, Bacterial (genetics), Deoxyribonucleases, Type II Site-Specific (metabolism), Genome, Bacterial (genetics), Genomic Library (MeSH), Humans (MeSH), Mycobacterium tuberculosis (classification), Mycobacterium tuberculosis (genetics), Polymerase Chain Reaction (MeSH), Sequence Analysis, DNA (MeSH), Transformation, Genetic (MeSH), Tuberculosis (microbiology).
- MESH :
- chemical , chemistry : DNA, Bacterial.
- chemical , genetics : DNA Transposable Elements, DNA, Bacterial.
- chemical , metabolism : Deoxyribonucleases, Type II Site-Specific.
- classification : Mycobacterium tuberculosis.
- genetics : Genome, Bacterial, Mycobacterium tuberculosis.
- methods : Chromosome Mapping.
- microbiology : Tuberculosis.
- Cloning, Molecular, Genomic Library, Humans, Polymerase Chain Reaction, Sequence Analysis, DNA, Transformation, Genetic.
Abstract
Evidence suggests that insertion of the IS6110 element is not without consequence to the biology of Mycobacterium tuberculosis complex strains. Thus, mapping of multiple IS6110 insertion sites in the genome of biomedically relevant clinical isolates would result in a better understanding of the role of this mobile element, particularly with regard to transmission, adaptability and virulence. In the present paper, we describe a versatile strategy, referred to as GL-PCR, that amplifies IS6110-flanking sequences based on the construction of a genomic library. M. tuberculosis chromosomal DNA is fully digested with HincII and then ligated into a plasmid vector between T7 and T3 promoter sequences. The ligation reaction product is transformed into Escherichia coli and selective PCR amplification targeting both 5' and 3' IS6110-flanking sequences are performed on the plasmid library DNA. For this purpose, four separate PCR reactions are performed, each combining an outward primer specific for one IS6110 end with either T7 or T3 primer. Determination of the nucleotide sequence of the PCR products generated from a single ligation reaction allowed mapping of 21 out of the 24 IS6110 copies of two 12 banded M. tuberculosis strains, yielding an overall sensitivity of 87,5%. Furthermore, by simply comparing the migration pattern of GL-PCR-generated products, the strategy proved to be as valuable as IS6110 RFLP for molecular typing of M. tuberculosis complex strains. Importantly, GL-PCR was able to discriminate between strains differing by a single IS6110 band.
DOI: 10.1016/j.mimet.2006.03.021
PubMed: 16725220
Affiliations:
Links toward previous steps (curation, corpus...)
- to stream PubMed, to step Corpus: 000796
- to stream PubMed, to step Curation: 000792
- to stream PubMed, to step Checkpoint: 000797
- to stream Main, to step Merge: 000029
- to stream Main, to step Curation: 000029
Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">A genomic library-based amplification approach (GL-PCR) for the mapping of multiple IS6110 insertion sites and strain differentiation of Mycobacterium tuberculosis.</title><author><name sortKey="Namouchi, Amine" sort="Namouchi, Amine" uniqKey="Namouchi A" first="Amine" last="Namouchi">Amine Namouchi</name><affiliation wicri:level="1"><nlm:affiliation>Laboratory of Mycobacteria, Institut Pasteur de Tunis, 13, Place Pasteur, BP 74, 1002, Tunis-Belvedre, Tunisie, Tunisia.</nlm:affiliation><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratory of Mycobacteria, Institut Pasteur de Tunis, 13, Place Pasteur, BP 74, 1002, Tunis-Belvedre, Tunisie</wicri:regionArea><wicri:noRegion>Tunisie</wicri:noRegion></affiliation></author><author><name sortKey="Mardassi, Helmi" sort="Mardassi, Helmi" uniqKey="Mardassi H" first="Helmi" last="Mardassi">Helmi Mardassi</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2006">2006</date><idno type="RBID">pubmed:16725220</idno><idno type="pmid">16725220</idno><idno type="doi">10.1016/j.mimet.2006.03.021</idno><idno type="wicri:Area/PubMed/Corpus">000796</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">000796</idno><idno type="wicri:Area/PubMed/Curation">000792</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">000792</idno><idno type="wicri:Area/PubMed/Checkpoint">000797</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">000797</idno><idno type="wicri:Area/Main/Merge">000029</idno><idno type="wicri:Area/Main/Curation">000029</idno><idno type="wicri:Area/Main/Exploration">000029</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">A genomic library-based amplification approach (GL-PCR) for the mapping of multiple IS6110 insertion sites and strain differentiation of Mycobacterium tuberculosis.</title><author><name sortKey="Namouchi, Amine" sort="Namouchi, Amine" uniqKey="Namouchi A" first="Amine" last="Namouchi">Amine Namouchi</name><affiliation wicri:level="1"><nlm:affiliation>Laboratory of Mycobacteria, Institut Pasteur de Tunis, 13, Place Pasteur, BP 74, 1002, Tunis-Belvedre, Tunisie, Tunisia.</nlm:affiliation><country xml:lang="fr">Tunisie</country><wicri:regionArea>Laboratory of Mycobacteria, Institut Pasteur de Tunis, 13, Place Pasteur, BP 74, 1002, Tunis-Belvedre, Tunisie</wicri:regionArea><wicri:noRegion>Tunisie</wicri:noRegion></affiliation></author><author><name sortKey="Mardassi, Helmi" sort="Mardassi, Helmi" uniqKey="Mardassi H" first="Helmi" last="Mardassi">Helmi Mardassi</name></author></analytic><series><title level="j">Journal of microbiological methods</title><idno type="ISSN">0167-7012</idno><imprint><date when="2006" type="published">2006</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Chromosome Mapping (methods)</term><term>Cloning, Molecular (MeSH)</term><term>DNA Transposable Elements (genetics)</term><term>DNA, Bacterial (chemistry)</term><term>DNA, Bacterial (genetics)</term><term>Deoxyribonucleases, Type II Site-Specific (metabolism)</term><term>Genome, Bacterial (genetics)</term><term>Genomic Library (MeSH)</term><term>Humans (MeSH)</term><term>Mycobacterium tuberculosis (classification)</term><term>Mycobacterium tuberculosis (genetics)</term><term>Polymerase Chain Reaction (MeSH)</term><term>Sequence Analysis, DNA (MeSH)</term><term>Transformation, Genetic (MeSH)</term><term>Tuberculosis (microbiology)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN bactérien (composition chimique)</term><term>ADN bactérien (génétique)</term><term>Analyse de séquence d'ADN (MeSH)</term><term>Banque génomique (MeSH)</term><term>Cartographie chromosomique (méthodes)</term><term>Clonage moléculaire (MeSH)</term><term>Génome bactérien (génétique)</term><term>Humains (MeSH)</term><term>Mycobacterium tuberculosis (classification)</term><term>Mycobacterium tuberculosis (génétique)</term><term>Réaction de polymérisation en chaîne (MeSH)</term><term>Transformation génétique (MeSH)</term><term>Tuberculose (microbiologie)</term><term>Type II site-specific deoxyribonuclease (métabolisme)</term><term>Éléments transposables d'ADN (génétique)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA Transposable Elements</term><term>DNA, Bacterial</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Deoxyribonucleases, Type II Site-Specific</term></keywords><keywords scheme="MESH" qualifier="classification" xml:lang="en"><term>Mycobacterium tuberculosis</term></keywords><keywords scheme="MESH" qualifier="composition chimique" xml:lang="fr"><term>ADN bactérien</term><term>Mycobacterium tuberculosis</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Genome, Bacterial</term><term>Mycobacterium tuberculosis</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ADN bactérien</term><term>Génome bactérien</term><term>Mycobacterium tuberculosis</term><term>Éléments transposables d'ADN</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Chromosome Mapping</term></keywords><keywords scheme="MESH" qualifier="microbiologie" xml:lang="fr"><term>Tuberculose</term></keywords><keywords scheme="MESH" qualifier="microbiology" xml:lang="en"><term>Tuberculosis</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Type II site-specific deoxyribonuclease</term></keywords><keywords scheme="MESH" qualifier="méthodes" xml:lang="fr"><term>Cartographie chromosomique</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Cloning, Molecular</term><term>Genomic Library</term><term>Humans</term><term>Polymerase Chain Reaction</term><term>Sequence Analysis, DNA</term><term>Transformation, Genetic</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Analyse de séquence d'ADN</term><term>Banque génomique</term><term>Clonage moléculaire</term><term>Humains</term><term>Réaction de polymérisation en chaîne</term><term>Transformation génétique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Evidence suggests that insertion of the IS6110 element is not without consequence to the biology of Mycobacterium tuberculosis complex strains. Thus, mapping of multiple IS6110 insertion sites in the genome of biomedically relevant clinical isolates would result in a better understanding of the role of this mobile element, particularly with regard to transmission, adaptability and virulence. In the present paper, we describe a versatile strategy, referred to as GL-PCR, that amplifies IS6110-flanking sequences based on the construction of a genomic library. M. tuberculosis chromosomal DNA is fully digested with HincII and then ligated into a plasmid vector between T7 and T3 promoter sequences. The ligation reaction product is transformed into Escherichia coli and selective PCR amplification targeting both 5' and 3' IS6110-flanking sequences are performed on the plasmid library DNA. For this purpose, four separate PCR reactions are performed, each combining an outward primer specific for one IS6110 end with either T7 or T3 primer. Determination of the nucleotide sequence of the PCR products generated from a single ligation reaction allowed mapping of 21 out of the 24 IS6110 copies of two 12 banded M. tuberculosis strains, yielding an overall sensitivity of 87,5%. Furthermore, by simply comparing the migration pattern of GL-PCR-generated products, the strategy proved to be as valuable as IS6110 RFLP for molecular typing of M. tuberculosis complex strains. Importantly, GL-PCR was able to discriminate between strains differing by a single IS6110 band.</div></front></TEI><affiliations><list><country><li>Tunisie</li></country></list><tree><noCountry><name sortKey="Mardassi, Helmi" sort="Mardassi, Helmi" uniqKey="Mardassi H" first="Helmi" last="Mardassi">Helmi Mardassi</name></noCountry><country name="Tunisie"><noRegion><name sortKey="Namouchi, Amine" sort="Namouchi, Amine" uniqKey="Namouchi A" first="Amine" last="Namouchi">Amine Namouchi</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Wicri/Sante/explor/MaghrebDataLibMedV2/Data/Main/Exploration
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 000029 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd -nk 000029 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Wicri/Sante
|area= MaghrebDataLibMedV2
|flux= Main
|étape= Exploration
|type= RBID
|clé= pubmed:16725220
|texte= A genomic library-based amplification approach (GL-PCR) for the mapping of multiple IS6110 insertion sites and strain differentiation of Mycobacterium tuberculosis.
}}
Pour générer des pages wiki
HfdIndexSelect -h $EXPLOR_AREA/Data/Main/Exploration/RBID.i -Sk "pubmed:16725220" \
| HfdSelect -Kh $EXPLOR_AREA/Data/Main/Exploration/biblio.hfd \
| NlmPubMed2Wicri -a MaghrebDataLibMedV2
| This area was generated with Dilib version V0.6.38. |  |