Incorporation of guanosine gels into sieving matrices for length- and sequence-based separation of DNA in capillary electrophoresis.
Identifieur interne : 002457 ( Main/Exploration ); précédent : 002456; suivant : 002458Incorporation of guanosine gels into sieving matrices for length- and sequence-based separation of DNA in capillary electrophoresis.
Auteurs : Yingying Dong [États-Unis] ; Linda B. McgownSource :
- Electrophoresis [ 1522-2683 ] ; 2011.
Descripteurs français
- KwdFr :
- ADN (), ADN (isolement et purification), Acrylamides (), Analyse de séquence d'ADN (), Conformation d'acide nucléique, Données de séquences moléculaires, Gels (), Guanosine (), Métagénome, Spectrométrie de masse MALDI, Séquence nucléotidique, Électrophorèse capillaire (), Électrophorèse capillaire (instrumentation).
- MESH :
English descriptors
- KwdEn :
- Acrylamides (chemistry), Base Sequence, DNA (chemistry), DNA (isolation & purification), Electrophoresis, Capillary (instrumentation), Electrophoresis, Capillary (methods), Gels (chemistry), Guanosine (chemistry), Metagenome, Molecular Sequence Data, Nucleic Acid Conformation, Sequence Analysis, DNA (methods), Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization.
- MESH :
- chemical , chemistry : Acrylamides, DNA, Gels, Guanosine.
- chemical , isolation & purification : DNA.
- instrumentation : Electrophoresis, Capillary.
- methods : Electrophoresis, Capillary, Sequence Analysis, DNA.
- Base Sequence, Metagenome, Molecular Sequence Data, Nucleic Acid Conformation, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization.
Abstract
Sieving gels are used in capillary gel electrophoresis to resolve DNA strands of different lengths. For complex samples, however, such as those encountered in metagenomic analysis of microbial communities or biofilms, length-based separation may mask the true genetic diversity of the community since different organisms may contribute same-length DNA with different sequences. There is a need, therefore, for DNA separations based on both the length and sequence. Previous work has demonstrated the ability of guanosine gels (G-gels) to separate four single-stranded DNA 76-mers that differ by only a few A/G base substitutions. The goal of the present work is to determine whether G-gels could be combined with commercial sieving gels in order to simultaneously separate DNA based on both length and sequence. The results are given for the four 76-mers and for a standard dsDNA ladder. Commercial sieving gels were used alone and in combination with G-gels. For the 76-mers, the combined medium was less efficient than the G-gel alone but was able to achieve partial resolution. The combined medium was at least as effective as the sieving gel alone at resolving the denatured DNA ladder and showed indications of sequence-based resolution as well, as supported by MALDI-MS. The results show that the combined sieving gel/G-gel medium retains the selectivity of the individual media, providing a promising approach to simultaneous length- and sequence-based DNA separation for metagenomic analysis of complex systems.
DOI: 10.1002/elps.201000545
PubMed: 21544840
Affiliations:
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Le document en format XML
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Mcgown</name></author></analytic><series><title level="j">Electrophoresis</title><idno type="eISSN">1522-2683</idno><imprint><date when="2011" type="published">2011</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Acrylamides (chemistry)</term><term>Base Sequence</term><term>DNA (chemistry)</term><term>DNA (isolation & purification)</term><term>Electrophoresis, Capillary (instrumentation)</term><term>Electrophoresis, Capillary (methods)</term><term>Gels (chemistry)</term><term>Guanosine (chemistry)</term><term>Metagenome</term><term>Molecular Sequence Data</term><term>Nucleic Acid Conformation</term><term>Sequence Analysis, DNA (methods)</term><term>Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN ()</term><term>ADN (isolement et purification)</term><term>Acrylamides ()</term><term>Analyse de séquence d'ADN ()</term><term>Conformation d'acide nucléique</term><term>Données de séquences moléculaires</term><term>Gels ()</term><term>Guanosine ()</term><term>Métagénome</term><term>Spectrométrie de masse MALDI</term><term>Séquence nucléotidique</term><term>Électrophorèse capillaire ()</term><term>Électrophorèse capillaire (instrumentation)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Acrylamides</term><term>DNA</term><term>Gels</term><term>Guanosine</term></keywords><keywords scheme="MESH" type="chemical" qualifier="isolation & purification" xml:lang="en"><term>DNA</term></keywords><keywords scheme="MESH" qualifier="instrumentation" xml:lang="en"><term>Electrophoresis, Capillary</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>ADN</term><term>Électrophorèse capillaire</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Electrophoresis, Capillary</term><term>Sequence Analysis, DNA</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Base Sequence</term><term>Metagenome</term><term>Molecular Sequence Data</term><term>Nucleic Acid Conformation</term><term>Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>ADN</term><term>Acrylamides</term><term>Analyse de séquence d'ADN</term><term>Conformation d'acide nucléique</term><term>Données de séquences moléculaires</term><term>Gels</term><term>Guanosine</term><term>Métagénome</term><term>Spectrométrie de masse MALDI</term><term>Séquence nucléotidique</term><term>Électrophorèse capillaire</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Sieving gels are used in capillary gel electrophoresis to resolve DNA strands of different lengths. For complex samples, however, such as those encountered in metagenomic analysis of microbial communities or biofilms, length-based separation may mask the true genetic diversity of the community since different organisms may contribute same-length DNA with different sequences. There is a need, therefore, for DNA separations based on both the length and sequence. Previous work has demonstrated the ability of guanosine gels (G-gels) to separate four single-stranded DNA 76-mers that differ by only a few A/G base substitutions. The goal of the present work is to determine whether G-gels could be combined with commercial sieving gels in order to simultaneously separate DNA based on both length and sequence. The results are given for the four 76-mers and for a standard dsDNA ladder. Commercial sieving gels were used alone and in combination with G-gels. For the 76-mers, the combined medium was less efficient than the G-gel alone but was able to achieve partial resolution. The combined medium was at least as effective as the sieving gel alone at resolving the denatured DNA ladder and showed indications of sequence-based resolution as well, as supported by MALDI-MS. The results show that the combined sieving gel/G-gel medium retains the selectivity of the individual media, providing a promising approach to simultaneous length- and sequence-based DNA separation for metagenomic analysis of complex systems.</div></front></TEI><affiliations><list><country><li>États-Unis</li></country><region><li>État de New York</li></region></list><tree><noCountry><name sortKey="Mcgown, Linda B" sort="Mcgown, Linda B" uniqKey="Mcgown L" first="Linda B" last="Mcgown">Linda B. Mcgown</name></noCountry><country name="États-Unis"><region name="État de New York"><name sortKey="Dong, Yingying" sort="Dong, Yingying" uniqKey="Dong Y" first="Yingying" last="Dong">Yingying Dong</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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