Ultrahigh-Throughput Multiplexing and Sequencing of >500-Base-Pair Amplicon Regions on the Illumina HiSeq 2500 Platform.
16S RNA
PCR
amplicon sequencing
ultrahigh throughput
Journal
mSystems
ISSN: 2379-5077
Titre abrégé: mSystems
Pays: United States
ID NLM: 101680636
Informations de publication
Date de publication:
Historique:
received:
18
01
2019
accepted:
23
01
2019
entrez:
26
2
2019
pubmed:
26
2
2019
medline:
26
2
2019
Statut:
epublish
Résumé
Amplification, sequencing, and analysis of the 16S rRNA gene affords characterization of microbial community composition. As this tool has become more popular and amplicon-sequencing applications have grown in the total number of samples, growth in sample multiplexing is becoming necessary while maintaining high sequence quality and sequencing depth. Here, modifications to the Illumina HiSeq 2500 platform are described which produce greater multiplexing capabilities and 300-bp paired-end reads of higher quality than those produced by the current Illumina MiSeq platform. To improve the feasibility and flexibility of this method, a 2-step PCR amplification protocol is also described that allows for targeting of different amplicon regions, and enhances amplification success from samples with low bacterial bioburden.
Identifiants
pubmed: 30801027
doi: 10.1128/mSystems.00029-19
pii: mSystems00029-19
pmc: PMC6381223
pii:
doi:
Types de publication
Journal Article
Langues
eng
Subventions
Organisme : NINR NIH HHS
ID : R01 NR015495
Pays : United States
Organisme : NIAID NIH HHS
ID : U19 AI084044
Pays : United States
Organisme : NIAID NIH HHS
ID : R01 AI116799
Pays : United States
Organisme : NIAID NIH HHS
ID : F32 AI136400
Pays : United States
Organisme : NIAID NIH HHS
ID : R01 AI089878
Pays : United States
Organisme : NIAID NIH HHS
ID : R21 AI107224
Pays : United States
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