scholarly journals Use of the DNA sequence of variable regions of the 16S rRNA gene for rapid and accurate identification of bacteria in theLactobacillus acidophiluscomplex

2000 ◽  
Vol 89 (3) ◽  
pp. 511-516 ◽  
Author(s):  
M.J. Kullen ◽  
R.B. Sanozky-Dawes ◽  
D.C. Crowell ◽  
T.R. Klaenhammer
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Sequence ◽  
Rrna Gene ◽  
Accurate Identification ◽  
Variable Regions ◽  
Identification Of Bacteria ◽  
The 16S Rrna Gene

Rapid identification of bacteria by real-time amplification and sequencing of the 16S rRNA gene

2006 ◽  
Vol 66 (1) ◽  
pp. 156-164 ◽  
Author(s):  
Inge Vliegen ◽  
Jan A. Jacobs ◽  
Erik Beuken ◽  
Cathrien A. Bruggeman ◽  
Cornelis Vink
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Real Time ◽  
Rapid Identification ◽  
Rrna Gene ◽  
Identification Of Bacteria ◽  
Time Amplification ◽  
The 16S Rrna Gene

scholarly journals Combining 16S rRNA gene variable regions enables high-resolution microbial community profiling

2017 ◽  
Author(s):  
Garold Fuks ◽  
Michael Elgart ◽  
Amnon Amir ◽  
Amit Zeisel ◽  
Peter J. Turnbaugh ◽  
...  
Keyword(s):  
High Resolution ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Read Length ◽  
Rrna Gene ◽  
Sample Number ◽  
Variable Regions ◽  
Total Length ◽  
Fragmented Dna ◽  
The 16S Rrna Gene

AbstractBackgroundMost of our knowledge about the remarkable microbial diversity on Earth comes from sequencing the 16S rRNA gene. The use of next-generation sequencing methods has increased sample number and sequencing depth, but the read length of the most widely used sequencing platforms today is quite short, requiring the researcher to choose a subset of the gene to sequence (typically 16-33% of the total length). Thus, many bacteria may share the same amplified region and the resolution of profiling is inherently limited. Platforms that offer ultra long read lengths, whole genome shotgun sequencing approaches, and computational frameworks formerly suggested by us and by others, all allow different ways to circumvent this problem yet suffer various shortcomings. There is need for a simple and low cost 16S rRNA gene based profiling approach that harnesses the short read length to provide a much larger coverage of the gene to allow for high resolution, even in harsh conditions of low bacterial biomass and fragmented DNA.ResultsThis manuscript suggests Short MUltiple Regions Framework (SMURF), a method to combine sequencing results from different PCR-amplified regions to provide one coherent profiling. The de facto amplicon length is the total length of all amplified regions, thus providing much higher resolution compared to current techniques. Computationally, the method solves a convex optimization problem that allows extremely fast reconstruction and requires only moderate memory. We demonstrate the increase in resolution by in silico simulations and by profiling two mock mixtures and real-world biological samples. Reanalyzing a mock mixture from the Human Microbiome Project achieved about two-fold improvement in resolution when combing two independent regions. Using a custom set of six primer pairs spanning about 1200bp (80%) of the 16S rRNA gene we were able to achieve ~100 fold improvement in resolution compared to a single region, over a mock mixture of common human gut bacterial isolates. Finally, profiling of a Drosophila melanogaster microbiome using the set of six primer pairs provided a ~100 fold increase in resolution, and thus enabling efficient downstream analysis.ConclusionsSMURF enables identification of near full-length 16S rRNA gene sequences in microbial communities, having resolution superior compared to current techniques. It may be applied to standard sample preparation protocols with very little modifications. SMURF also paves the way to high-resolution profiling of low-biomass and fragmented DNA, e.g., in the case of Formalin-fixed and Paraffin-embedded samples, fossil-derived DNA or DNA exposed to other degrading conditions. The approach is not restricted to combining amplicons of the 16S rRNA gene and may be applied to any set of amplicons, e.g., in Multilocus Sequence Typing (MLST).

scholarly journals Resolving microbial microdiversity with high accuracy full length 16S rRNA Illumina sequencing

2014 ◽  
Author(s):  
Catherine Burke ◽  
Aaron E Darling
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Single Molecule ◽  
Illumina Miseq ◽  
Full Length ◽  
Rrna Gene ◽  
Variable Regions ◽  
Phylogenetic Resolution ◽  
Miseq Platform ◽  
The 16S Rrna Gene

We describe a method for sequencing full-length 16S rRNA gene amplicons using the high throughput Illumina MiSeq platform. The resulting sequences have about 100-fold higher accuracy than standard Illumina reads and are chimera filtered using information from a single molecule dual tagging scheme that boosts the signal available for chimera detection. We demonstrate that the data provides fine scale phylogenetic resolution not available from Illumina amplicon methods targeting smaller variable regions of the 16S rRNA gene.

scholarly journals PEMANFAATAN GEN 16S rRNA SEBAGAI PERANGKAT IDENTIFIKASI BAKTERI UNTUK PENELITIAN-PENELITIAN DI INDONESIA

PHARMACON ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 16
Author(s):  
Claudia Valleria Akihary ◽  
Beivy Jonathan Kolondam
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Species ◽  
Variable Region ◽  
Animal Husbandry ◽  
Agricultural Science ◽  
Research Tool ◽  
Rrna Gene ◽  
Accurate Identification ◽  
Identification Of Bacteria

ABSTRACTThe 16S rRNA gene has hyper variable region and different for one bacterial species to another. The gene is being used as research tool to help for accurate identification of bacteria in many fields in Indonesia. As a useful tool, the 16S rRNA gene sequence is important as to explore the potencies of a bacterial species. Sequencing of this gene is very useful for research in clinical study, fisheries, marine science, agricultural science, and animal husbandry in Indonesia.   Keywords: 16S rRNA gene, research tool, bacteria, Indonesia                                                                     ABSTRAKGen 16S rRNA memiliki region yang sangat bervariasi dan berbeda setiap spesies bakteri. Penggunaannya sebagai perangkat penelitian, gen 16S rRNA telah banyak membantu dalam proses identifikasi berbagai jenis bakteri secara akurat untuk berbagai penelitian di Indonesia. Gen 16S rRNA tidak hanya dapat mengidentifikasi tetapi dapat dijadikan arahan dalam mengetahui potensi suatu bakteri. Sekuensing gen 16S rRNA telah digunakan secara luas untuk penelitian di bidang klinis, perikanan, kelautan, pertanian dan peternakan di Indonesia.Kata Kunci: Gen 16S rRNA, perangkat penelitian, Bakteri, Indonesia.

scholarly journals Molecular identification of bacteria isolated from culture medium of rotifer fed on fishery waste diet

2020 ◽  
Vol 21 (6) ◽  
Author(s):  
Stenly Wullur ◽  
HATOPAN NAPITUPULU ◽  
LETHA LOISE WANTANIA ◽  
ELVY LIKE GINTING ◽  
VEIBE WAROUW ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Molecular Identification ◽  
Culture Medium ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Seawater Salinity ◽  
Identification Of Bacteria ◽  
The 16S Rrna Gene

Abstract. Wullur S, Napitupulu H, Wantania LL, Ginting EL, Warouw V, Tallei TE, Rumengan IFM. 2020. Molecular identification of bacteria isolated from culture medium of rotifer fed on fishery waste diet. Biodiversitas 21: 2735-2740. The aim of this study was 16S-rRNA sequences based molecular identification of bacteria isolated from culture medium of rotifer fed with fishery waste diet (FWD). We cultured rotifer Brachionus rotundiformis in sterilized seawater (salinity 25 ppt) using FWD, following the procedure in Patent No. P00201609066. Bacteria from the culture were collected, homogenized, diluted 10 to 1000 fold, spread on agar plates and incubated at 370C for 24 to 48 hours. Representative colonies of the bacteria according to their morphologies were isolated for further characterization. Genomic DNA of the isolates were extracted, and the 16S rRNA gene of the isolates were amplified. Polymerase Chain Reaction (PCR) product of each isolate was sequenced and queried against the NCBI GenBank database. Six different isolates based on size, color, elevation, margin, and colony were observed during 24-48 hours incubation at 370C. The 16S rRNA genes of the six isolates were successfully amplified and produced DNA band at 1300-1500 bp, with quality value equal to or greater than 20 (QV20+) of each entire sequence around 941-1253 bases. Basic Local Alignment Search Tool (BLAST) queries in the NCBI GenBank and EzBioCloud database using the 16S-rRNA gene sequences showed that the six isolates belong to four different genera, i.e: Bacillus, Staphylococcus, Vibrio, and Alteromonas.

scholarly journals Limitations of 16S rRNA Gene Sequencing to Characterize Lactobacillus Species in the Upper Genital Tract

2021 ◽  
Vol 9 ◽  
Author(s):  
Jessica L. O’Callaghan ◽  
Dana Willner ◽  
Melissa Buttini ◽  
Flavia Huygens ◽  
Elise S. Pelzer
Keyword(s):  
Microbial Community ◽  
Next Generation Sequencing ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Genital Tract ◽  
Rrna Gene ◽  
Next Generation ◽  
Variable Regions ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

The endometrial cavity is an upper genital tract site previously thought as sterile, however, advances in culture-independent, next-generation sequencing technology have revealed that this low-biomass site harbors a rich microbial community which includes multiple Lactobacillus species. These bacteria are considered to be the most abundant non-pathogenic genital tract commensals. Next-generation sequencing of the female lower genital tract has revealed significant variation amongst microbial community composition with respect to Lactobacillus sp. in samples collected from healthy women and women with urogenital conditions. The aim of this study was to evaluate our ability to characterize members of the genital tract microbial community to species-level taxonomy using variable regions of the 16S rRNA gene. Samples were interrogated for the presence of microbial DNA using next-generation sequencing technology that targets the V5–V8 regions of the 16S rRNA gene and compared to speciation using qPCR. We also performed re-analysis of published data using alternate variable regions of the 16S rRNA gene. In this analysis, we explore next-generation sequencing of clinical genital tract isolates as a method for high throughput identification to species-level of key Lactobacillus sp. Data revealed that characterization of genital tract taxa is hindered by a lack of a consensus protocol and 16S rRNA gene region target allowing comparison between studies.

scholarly journals Comparison of Two PCRs for Detection of Mycobacterium ulcerans

1999 ◽  
Vol 37 (1) ◽  
pp. 206-208 ◽  
Author(s):  
A. Guimaraes-Peres ◽  
F. Portaels ◽  
P. de Rijk ◽  
K. Fissette ◽  
S. R. Pattyn ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Sequence ◽  
Repetitive Dna ◽  
Repetitive Sequence ◽  
Mycobacterium Ulcerans ◽  
Rrna Gene ◽  
Clinical Specimens ◽  
Gene Coding ◽  
The 16S Rrna Gene

Two nested PCRs for the detection of Mycobacterium ulcerans were compared by using a collection of 65 clinical specimens. The first method amplifies the gene coding for 16S rRNA, and the second method amplifies a repetitive DNA sequence. The sensitivities of bacterioscopy, culture, 16S rRNA gene PCR, and repetitive-sequence PCR were 29, 34, 80, and 85%, respectively. Compared to the 16S rRNA gene PCR, the repetitive-sequence PCR was faster, easier to perform, and less expensive.

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