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16s rRNA Sequencing with MR DNA

16S ribosomal  (rRNA) sequencing using next generation sequencing is a method used to identify and compare bacteria and archaea present within almost any type of sample. 16S rRNA gene sequencing is a well-established method for studying phylogeny and taxonomy of samples from complex microbiomes or environments that are difficult or impossible to study.





16s sequencing illumina or PGM low cost prices with MR DNA

MR DNA is a next generation sequencing provider with low cost 16s sequencing services.




248. Gut Microbes. 2012 Nov-Dec;3(6):572-6. doi: 10.4161/gmic.21772. Epub 2012 Aug 16.


The microbiota link to irritable bowel syndrome: an emerging story.


Jeffery IB(1), Quigley EM, Öhman L, Simrén M, O'Toole PW.


Author information:

(1)Department of Microbiology, University College Cork, Cork, Ireland.


Irritable Bowel Syndrome (IBS) is a clinically heterogeneous disorder which is

likely to involve a number of causative factors. The contribution of altered

intestinal microbiota composition or function to this disorder is controversial,

and is the subject of much current research. Until recently, the technical

limitations of the methodologies available have not permitted an adequate survey

of low-abundance microbial species. Recent technological developments have

enabled the analysis of the global population of the microbiome using high

through-put, culture independent, 16S rRNA amplicon pyrosequencing. Using these

new methodologies, we are able to gain important biological insights into the

link between functional bowel disorders and the microbiome. This addendum

contextualizes and summarizes the results of these studies, and defines the

future challenges and opportunities in the field.


DOI: 10.4161/gmic.21772

PMCID: PMC3495796

PMID: 22895081  [PubMed - indexed for MEDLINE]



249. Pathol Biol (Paris). 2012 Jun;60(3):e30-5. doi: 10.1016/j.patbio.2011.03.014.

Epub 2011 May 31.


Intragenomic and intraspecific heterogeneity of the 16S rRNA gene in seven

bacterial species from the respiratory tract of cystic fibrosis patients assessed

by PCR-Temporal Temperature Gel Electrophoresis.


Michon AL(1), Jumas-Bilak E, Imbert A, Aleyrangues L, Counil F, Chiron R,

Marchandin H.


Author information:

(1)UMR 5119 équipe pathogènes et environnements, UFR des sciences pharmaceutiques

et biologiques, faculté de pharmacie, université Montpellier 1, 15, avenue

Charles-Flahault, BP 14491, 34093 Montpellier cedex 5, France.


16S rRNA gene-based cultivation-independent methods are increasingly used to

study the diversity of microbiota during health and disease. One bias of these

methods is the variability of 16S rRNA gene that may exist among strains of a

same species (intraspecific heterogeneity) or between rrs copies in a genome

(intragenomic heterogeneity). We evaluated the level of intraspecific and

intragenomic 16S rDNA variability in seven species frequently encountered in

respiratory tract samples in cystic fibrosis (CF). A total of 179 strains were

subjected to V3 region 16S rDNA PCR-TTGE. Using this easy-to-perform and rapid

method, different levels of V3 region rrs heterogeneity were demonstrated. No

intraspecific and intragenomic rrs heterogeneity was demonstrated for Moraxella

catarrhalis (n=16), Pseudomonas aeruginosa (n=31) and Streptococcus pneumoniae

(n=14) showing a single PCR-TTGE band characteristic of the species. Low level of

intraspecific heterogeneity was observed for Staphylococcus aureus (n=30),

Stenotrophomonas maltophilia (n=29) and Achromobacter xylosoxidans (n=28), and

17%, 38% and 96% of these strains showed intragenomic heterogeneity (two to four

different rrs copies), respectively. Haemophilus influenzae (n=31) displayed the

higher level of intraspecific variability with 23 different PCR-TTGE patterns and

61% of the strains showed intragenomic rrs heterogeneity (two to four different

rrs copies). Although only one hypervariable region of the 16S rRNA gene was

explored, intraspecific and intragenomic rrs heterogeneity was frequently

observed in this study and should be taken into consideration for a better

interpretation of 16S rRNA gene-based diversity profiles in denaturing gels and

to avoid any overestimation of the respiratory microbiota diversity in CF.


Copyright © 2011 Elsevier Masson SAS. All rights reserved.


DOI: 10.1016/j.patbio.2011.03.014

PMID: 21621347  [PubMed - indexed for MEDLINE]



250. PLoS One. 2012;7(8):e43052. doi: 10.1371/journal.pone.0043052. Epub 2012 Aug 15.


Analysis of the gut microbiota in the old order Amish and its relation to the

metabolic syndrome.


Zupancic ML(1), Cantarel BL, Liu Z, Drabek EF, Ryan KA, Cirimotich S, Jones C,

Knight R, Walters WA, Knights D, Mongodin EF, Horenstein RB, Mitchell BD, Steinle

N, Snitker S, Shuldiner AR, Fraser CM.


Author information:

(1)Institute for Genome Sciences, University of Maryland School of Medicine,

Baltimore, Maryland, United States of America.


Obesity has been linked to the human gut microbiota; however, the contribution of

gut bacterial species to the obese phenotype remains controversial because of

conflicting results from studies in different populations. To explore the

possible dysbiosis of gut microbiota in obesity and its metabolic complications,

we studied men and women over a range of body mass indices from the Old Order

Amish sect, a culturally homogeneous Caucasian population of Central European

ancestry. We characterized the gut microbiota in 310 subjects by deep

pyrosequencing of bar-coded PCR amplicons from the V1-V3 region of the 16S rRNA

gene. Three communities of interacting bacteria were identified in the gut

microbiota, analogous to previously identified gut enterotypes. Neither BMI nor

any metabolic syndrome trait was associated with a particular gut community.

Network analysis identified twenty-two bacterial species and four OTUs that were

either positively or inversely correlated with metabolic syndrome traits,

suggesting that certain members of the gut microbiota may play a role in these

metabolic derangements.


DOI: 10.1371/journal.pone.0043052

PMCID: PMC3419686

PMID: 22905200  [PubMed - indexed for MEDLINE]



251. PLoS One. 2010 May 17;5(5):e10667. doi: 10.1371/journal.pone.0010667.


Through ageing, and beyond: gut microbiota and inflammatory status in seniors and



Biagi E(1), Nylund L, Candela M, Ostan R, Bucci L, Pini E, Nikkïla J, Monti D,

Satokari R, Franceschi C, Brigidi P, De Vos W.


Author information:

(1)Department of Pharmaceutical Sciences, University of Bologna, Bologna, Italy.


Erratum in

    PLoS One. 2010;5(6). doi:



BACKGROUND: Age-related physiological changes in the gastrointestinal tract, as

well as modifications in lifestyle, nutritional behaviour, and functionality of

the host immune system, inevitably affect the gut microbiota, resulting in a

greater susceptibility to infections.

METHODOLOGY/PRINCIPAL FINDINGS: By using the Human Intestinal Tract Chip

(HITChip) and quantitative PCR of 16S rRNA genes of Bacteria and Archaea, we

explored the age-related differences in the gut microbiota composition among

young adults, elderly, and centenarians, i.e subjects who reached the extreme

limits of the human lifespan, living for over 100 years. We observed that the

microbial composition and diversity of the gut ecosystem of young adults and

seventy-years old people is highly similar but differs significantly from that of

the centenarians. After 100 years of symbiotic association with the human host,

the microbiota is characterized by a rearrangement in the Firmicutes population

and an enrichment in facultative anaerobes, notably pathobionts. The presence of

such a compromised microbiota in the centenarians is associated with an increased

inflammatory status, also known as inflammageing, as determined by a range of

peripheral blood inflammatory markers. This may be explained by a remodelling of

the centenarians' microbiota, with a marked decrease in Faecalibacterium

prauznitzii and relatives, symbiotic species with reported anti-inflammatory

properties. As signature bacteria of the long life we identified specifically

Eubacterium limosum and relatives that were more than ten-fold increased in the


CONCLUSIONS/SIGNIFICANCE: We provide evidence for the fact that the ageing

process deeply affects the structure of the human gut microbiota, as well as its

homeostasis with the host's immune system. Because of its crucial role in the

host physiology and health status, age-related differences in the gut microbiota

composition may be related to the progression of diseases and frailty in the

elderly population.


DOI: 10.1371/journal.pone.0010667

PMCID: PMC2871786

PMID: 20498852  [PubMed - indexed for MEDLINE]



252. Appl Microbiol Biotechnol. 2014 Apr;98(7):3317-26. doi:

10.1007/s00253-013-5402-z. Epub 2013 Dec 5.


Tracking human sewage microbiome in a municipal wastewater treatment plant.


Cai L(1), Ju F, Zhang T.


Author information:

(1)Environmental Biotechnology Laboratory, Department of Civil Engineering, The

University of Hong Kong, Hong Kong, SAR, China.


Human sewage pollution is a major threat to public health because sewage always

comes with pathogens. Human sewage is usually received and treated by wastewater

treatment plants (WWTPs) to control pathogenic risks and ameliorate environmental

health. However, untreated sewage that flows into water environments may cause

serious waterborne diseases, as reported in India and Bangladesh. To examine the

fate of the human sewage microbiome in a local municipal WWTP of Hong Kong, we

used massively parallel sequencing of 16S rRNA gene to systematically profile

microbial communities in samples from three sections (i.e., influent, activated

sludge, and effluent) obtained monthly throughout 1 year. The results indicated

that: (1) influent sewage bacterial profile reflected the human microbiome; (2)

human gut bacterial community was the dominant force shaping influent sewage

bacterial profile; (3) most human sewage bacteria could be effectively removed by

the WWTP; (4) a total of 75 genera were profiled as potentially pathogenic

bacteria, most of which were still present in the effluent although at a very low

level; (5) a grouped pattern of bacterial community was observed among the same

section samples but a dispersed pattern was found among the different section

samples; and (6) activated sludge was less affected by the influent sewage

bacteria, but it showed a significant impact on the effluent bacteria. All of

these findings provide novel insights toward a mechanistic understanding of the

fate of human sewage microbiome in the WWTP.


DOI: 10.1007/s00253-013-5402-z

PMID: 24305737  [PubMed - indexed for MEDLINE]



253. PLoS One. 2016 Feb 12;11(2):e0149095. doi: 10.1371/journal.pone.0149095.

eCollection 2016.


Metagenomic Sequencing of the Chronic Obstructive Pulmonary Disease Upper

Bronchial Tract Microbiome Reveals Functional Changes Associated with Disease



Cameron SJ(1), Lewis KE(2,)(3), Huws SA(1), Lin W(1), Hegarty MJ(1), Lewis PD(3),

Mur LA(1), Pachebat JA(1).


Author information:

(1)Institute of Biological, Environmental and Rural Sciences, Edward Llywd

Building, Penglais Campus, Aberystwyth, Ceredigion, SY23 3FG, United Kingdom.

(2)Department of Respiratory Medicine, Prince Phillip Hospital, Hywel Dda

University Health Board, Llanelli, SA14 8QF, United Kingdom. (3)College of

Medicine, Swansea University, Swansea, SA2 8PP, United Kingdom.


Chronic Obstructive Pulmonary Disease (COPD) is a major source of mortality and

morbidity worldwide. The microbiome associated with this disease may be an

important component of the disease, though studies to date have been based on

sequencing of the 16S rRNA gene, and have revealed unequivocal results. Here, we

employed metagenomic sequencing of the upper bronchial tract (UBT) microbiome to

allow for greater elucidation of its taxonomic composition, and revealing

functional changes associated with the disease. The bacterial metagenomes within

sputum samples from eight COPD patients and ten 'healthy' smokers (Controls) were

sequenced, and suggested significant changes in the abundance of bacterial

species, particularly within the Streptococcus genus. The functional capacity of

the COPD UBT microbiome indicated an increased capacity for bacterial growth,

which could be an important feature in bacterial-associated acute exacerbations.

Regression analyses correlated COPD severity (FEV1% of predicted) with

differences in the abundance of Streptococcus pneumoniae and functional

classifications related to a reduced capacity for bacterial sialic acid

metabolism. This study suggests that the COPD UBT microbiome could be used in

patient risk stratification and in identifying novel monitoring and treatment

methods, but study of a longitudinal cohort will be required to unequivocally

relate these features of the microbiome with COPD severity.


DOI: 10.1371/journal.pone.0149095

PMCID: PMC4752236

PMID: 26872143  [PubMed - indexed for MEDLINE]



254. Genome Med. 2012 Oct 10;4(10):77. doi: 10.1186/gm378. eCollection 2012.


Shifts in human skin and nares microbiota of healthy children and adults.


Oh J(1), Conlan S(1), Polley EC(2), Segre JA(1), Kong HH(3).


Author information:

(1)Genetics and Molecular Biology Branch, National Human Genome Research

Institute, NIH, 49 Convent Dr., Bethesda, MD 20814, USA. (2)Biometric Research

Branch, Division of Cancer Treatment and Diagnosis, National Cancer Institute,

NIH, 6130 Executive Blvd, Rockville, MD 20852, USA. (3)Dermatology Branch, Center

for Cancer Research, National Cancer Institute, NIH, 10 Center Dr., Bethesda, MD

20814, USA.


BACKGROUND: Characterization of the topographical and temporal diversity of the

microbial collective (microbiome) hosted by healthy human skin established a

reference for studying disease-causing microbiomes. Physiologic changes occur in

the skin as humans mature from infancy to adulthood. Thus, characterizations of

adult microbiomes might have limitations when considering pediatric disorders

such as atopic dermatitis (AD) or issues such as sites of microbial carriage. The

objective of this study was to determine if microbial communities at several body

sites in children differed significantly from adults.

METHODS: Using 16S-rRNA gene sequencing technology, we characterized and compared

the bacterial communities of four body sites in relation to Tanner stage of human

development. Body sites sampled included skin sites characteristically involved

in AD (antecubital/popliteal fossae), a control skin site (volar forearm), and

the nares. Twenty-eight healthy individuals aged from 2 to 40 years were

evaluated at the outpatient dermatology clinic in the National Institutes of

Health's Clinical Center. Exclusion criteria included the use of systemic

antibiotics within 6 months, current/prior chronic skin disorders, asthma,

allergic rhinitis, or other chronic medical conditions.

RESULTS: Bacterial communities in the nares of children (Tanner developmental

stage 1) differed strikingly from adults (Tanner developmental stage 5).

Firmicutes (Streptococcaceae), Bacteroidetes, and Proteobacteria (β, γ) were

overrepresented in Tanner 1 compared to Tanner 5 individuals, where

Corynebacteriaceae and Propionibacteriaceae predominated. While bacterial

communities were significantly different between the two groups in all sites, the

most marked microbial shifts were observed in the nares, a site that can harbor

pathogenic species, including Staphylococcus aureus and Streptococcus pneumonia.

CONCLUSIONS: Significant shifts in the microbiota associated with progressive

sexual maturation as measured by Tanner staging suggest that puberty-dependent

shifts in the skin and nares microbiomes may have significant implications

regarding prevention and treatment of pediatric disorders involving microbial

pathogens and colonization.


DOI: 10.1186/gm378

PMCID: PMC3580446

PMID: 23050952  [PubMed]





71. ISME J. 2012 Jan;6(1):183-94. doi: 10.1038/ismej.2011.74. Epub 2011 Jun 16.


Illumina-based analysis of microbial community diversity.


Degnan PH(1), Ochman H.


Author information:

(1)Department of Ecology and Evolutionary Biology, Yale University, New Haven,



Microbes commonly exist in milieus of varying complexity and diversity. Although

cultivation-based techniques have been unable to accurately capture the true

diversity within microbial communities, these deficiencies have been overcome by

applying molecular approaches that target the universally conserved 16S ribosomal

RNA gene. The recent application of 454 pyrosequencing to simultaneously sequence

thousands of 16S rDNA sequences (pyrotags) has revolutionized the

characterization of complex microbial communities. To date, studies based on 454

pyrotags have dominated the field, but sequencing platforms that generate many

more sequence reads at much lower costs have been developed. Here, we use the

Illumina sequencing platform to design a strategy for 16S amplicon analysis

(iTags), and assess its generality, practicality and potential complications. We

fabricated and sequenced paired-end libraries of amplified hyper-variable 16S

rDNA fragments from sets of samples that varied in their contents, ranging from a

single bacterium to highly complex communities. We adopted an approach that

allowed us to evaluate several potential sources of errors, including sequencing

artifacts, amplification biases, non-corresponding paired-end reads and mistakes

in taxonomic classification. By considering each source of error, we delineate

ways to make biologically relevant and robust conclusions from the millions of

sequencing reads that can be readily generated by this technology.


DOI: 10.1038/ismej.2011.74

PMCID: PMC3246231

PMID: 21677692  [PubMed - indexed for MEDLINE]



72. Syst Biol Reprod Med. 2011 Jun;57(3):162-70. doi: 10.3109/19396368.2011.555598.

Epub 2011 Mar 1.


Strategy for microbiome analysis using 16S rRNA gene sequence analysis on the

Illumina sequencing platform.


Ram JL(1), Karim AS, Sendler ED, Kato I.


Author information:

(1)Department of Physiology, Wayne State University, Detroit, MI 48201 USA.


Comment in

    Syst Biol Reprod Med. 2011 Jun;57(3):117-8.


Understanding the identity and changes of organisms in the urogenital and other

microbiomes of the human body may be key to discovering causes and new treatments

of many ailments, such as vaginosis. High-throughput sequencing technologies have

recently enabled discovery of the great diversity of the human microbiome. The

cost per base of many of these sequencing platforms remains high (thousands of

dollars per sample); however, the Illumina Genome Analyzer (IGA) is estimated to

have a cost per base less than one-fifth of its nearest competitor. The main

disadvantage of the IGA for sequencing PCR-amplified 16S rRNA genes is that the

maximum read-length of the IGA is only 100 bases; whereas, at least 300 bases are

needed to obtain phylogenetically informative data down to the genus and species

level. In this paper we describe and conduct a pilot test of a multiplex

sequencing strategy suitable for achieving total reads of > 300 bases per

extracted DNA molecule on the IGA. Results show that all proposed primers produce

products of the expected size and that correct sequences can be obtained, with

all proposed forward primers. Various bioinformatic optimization of the Illumina

Bustard analysis pipeline proved necessary to extract the correct sequence from

IGA image data, and these modifications of the data files indicate that further

optimization of the analysis pipeline may improve the quality rankings of the

data and enable more sequence to be correctly analyzed. The successful

application of this method could result in an unprecedentedly deep description

(800,000 taxonomic identifications per sample) of the urogenital and other

microbiomes in a large number of samples at a reasonable cost per sample.


DOI: 10.3109/19396368.2011.555598

PMID: 21361774  [PubMed - indexed for MEDLINE]



73. ISME J. 2011 Apr;5(4):741-9. doi: 10.1038/ismej.2010.160. Epub 2010 Oct 21.


BIPES, a cost-effective high-throughput method for assessing microbial diversity.


Zhou HW(1), Li DF, Tam NF, Jiang XT, Zhang H, Sheng HF, Qin J, Liu X, Zou F.


Author information:

(1)Department of Environmental Health, School of Public Health and Tropical

Medicine, Southern Medical University, Guangzhou, Guangdong, China.


Pyrosequencing of 16S rRNA (16S) variable tags has become the most popular method

for assessing microbial diversity, but the method remains costly for the

evaluation of large numbers of environmental samples with high sequencing depths.

We developed a barcoded Illumina paired-end (PE) sequencing (BIPES) method that

sequences each 16S V6 tag from both ends on the Illumina HiSeq 2000, and the PE

reads are then overlapped to obtain the V6 tag. The average accuracy of Illumina

single-end (SE) reads was only 97.9%, which decreased from ∼99.9% at the start of

the read to less than 85% at the end of the read; nevertheless, overlapping of

the PE reads significantly increased the sequencing accuracy to 99.65% by

verifying the 3' end of each SE in which the sequencing quality was degraded.

After the removal of tags with two or more mismatches within the medial 40-70

bases of the reads and of tags with any primer errors, the overall base

sequencing accuracy of the BIPES reads was further increased to 99.93%. The BIPES

reads reflected the amounts of the various tags in the initial template, but long

tags and high GC tags were underestimated. The BIPES method yields 20-50 times

more 16S V6 tags than does pyrosequencing in a single-flow cell run, and each of

the BIPES reads costs less than 1/40 of a pyrosequencing read. As a laborsaving

and cost-effective method, BIPES can be routinely used to analyze the microbial

ecology of both environmental and human microbiomes.


DOI: 10.1038/ismej.2010.160

PMCID: PMC3105743

PMID: 20962877  [PubMed - indexed for MEDLINE]



74. Nucleic Acids Res. 2010 Dec;38(22):e200. doi: 10.1093/nar/gkq873. Epub 2010 Sep



Comparison of two next-generation sequencing technologies for resolving highly

complex microbiota composition using tandem variable 16S rRNA gene regions.


Claesson MJ(1), Wang Q, O'Sullivan O, Greene-Diniz R, Cole JR, Ross RP, O'Toole



Author information:

(1)Department of Microbiology, University College Cork, Cork, Ireland.


High-throughput molecular technologies can profile microbial communities at high

resolution even in complex environments like the intestinal microbiota. Recent

improvements in next-generation sequencing technologies allow for even finer

resolution. We compared phylogenetic profiling of both longer (454 Titanium)

sequence reads with shorter, but more numerous, paired-end reads (Illumina). For

both approaches, we targeted six tandem combinations of 16S rRNA gene variable

regions, in microbial DNA extracted from a human faecal sample, in order to

investigate their limitations and potentials. In silico evaluations predicted

that the V3/V4 and V4/V5 regions would provide the highest classification

accuracies for both technologies. However, experimental sequencing of the V3/V4

region revealed significant amplification bias compared to the other regions,

emphasising the necessity for experimental validation of primer pairs. The latest

developments of 454 and Illumina technologies offered higher resolution compared

to their previous versions, and showed relative consistency with each other.

However, the majority of the Illumina reads could not be classified down to genus

level due to their shorter length and higher error rates beyond 60 nt.

Nonetheless, with improved quality and longer reads, the far greater coverage of

Illumina promises unparalleled insights into highly diverse and complex

environments such as the human gut.


DOI: 10.1093/nar/gkq873

PMCID: PMC3001100

PMID: 20880993  [PubMed - indexed for MEDLINE]



75. PLoS One. 2010 Aug 12;5(8):e12078. doi: 10.1371/journal.pone.0012078.


Deep sequencing of the vaginal microbiota of women with HIV.


Hummelen R(1), Fernandes AD, Macklaim JM, Dickson RJ, Changalucha J, Gloor GB,

Reid G.


Author information:

(1)Canadian Research & Development Centre for Probiotics, Lawson Health Research

Institute, London, Canada.


BACKGROUND: Women living with HIV and co-infected with bacterial vaginosis (BV)

are at higher risk for transmitting HIV to a partner or newborn. It is poorly

understood which bacterial communities constitute BV or the normal vaginal

microbiota among this population and how the microbiota associated with BV

responds to antibiotic treatment.

METHODS AND FINDINGS: The vaginal microbiota of 132 HIV positive Tanzanian women,

including 39 who received metronidazole treatment for BV, were profiled using

Illumina to sequence the V6 region of the 16S rRNA gene. Of note, Gardnerella

vaginalis and Lactobacillus iners were detected in each sample constituting core

members of the vaginal microbiota. Eight major clusters were detected with

relatively uniform microbiota compositions. Two clusters dominated by L. iners or

L. crispatus were strongly associated with a normal microbiota. The L. crispatus

dominated microbiota were associated with low pH, but when L. crispatus was not

present, a large fraction of L. iners was required to predict a low pH. Four

clusters were strongly associated with BV, and were dominated by Prevotella

bivia, Lachnospiraceae, or a mixture of different species. Metronidazole

treatment reduced the microbial diversity and perturbed the BV-associated

microbiota, but rarely resulted in the establishment of a lactobacilli-dominated


CONCLUSIONS: Illumina based microbial profiling enabled high though-put analyses

of microbial samples at a high phylogenetic resolution. The vaginal microbiota

among women living with HIV in Sub-Saharan Africa constitutes several profiles

associated with a normal microbiota or BV. Recurrence of BV frequently

constitutes a different BV-associated profile than before antibiotic treatment.


DOI: 10.1371/journal.pone.0012078

PMCID: PMC2920804

PMID: 20711427  [PubMed - indexed for MEDLINE]



76. J Microbiol Methods. 2009 Dec;79(3):266-71. doi: 10.1016/j.mimet.2009.09.012.

Epub 2009 Sep 29.


Metagenomic study of the oral microbiota by Illumina high-throughput sequencing.


Lazarevic V(1), Whiteson K, Huse S, Hernandez D, Farinelli L, Osterås M,

Schrenzel J, François P.


Author information:

(1)Genomic Research Laboratory, Geneva University Hospitals, Rue

Gabrielle-Perret-Gentil 4, CH-1211 Geneva 14, Switzerland.


To date, metagenomic studies have relied on the utilization and analysis of reads

obtained using 454 pyrosequencing to replace conventional Sanger sequencing.

After extensively scanning the 16S ribosomal RNA (rRNA) gene, we identified the

V5 hypervariable region as a short region providing reliable identification of

bacterial sequences available in public databases such as the Human Oral

Microbiome Database. We amplified samples from the oral cavity of three healthy

individuals using primers covering an approximately 82-base segment of the V5

loop, and sequenced using the Illumina technology in a single orientation. We

identified 135 genera or higher taxonomic ranks from the resulting 1,373,824

sequences. While the abundances of the most common phyla (Firmicutes,

Proteobacteria, Actinobacteria, Fusobacteria and TM7) are largely comparable to

previous studies, Bacteroidetes were less present. Potential sources for this

difference include classification bias in this region of the 16S rRNA gene, human

sample variation, sample preparation and primer bias. Using an Illumina

sequencing approach, we achieved a much greater depth of coverage than previous

oral microbiota studies, allowing us to identify several taxa not yet discovered

in these types of samples, and to assess that at least 30,000 additional reads

would be required to identify only one additional phylotype. The evolution of

high-throughput sequencing technologies, and their subsequent improvements in

read length enable the utilization of different platforms for studying

communities of complex flora. Access to large amounts of data is already leading

to a better representation of sample diversity at a reasonable cost.


DOI: 10.1016/j.mimet.2009.09.012

PMCID: PMC3568755

PMID: 19796657  [PubMed - indexed for MEDLINE]



16s rRNA Sequencing with MR DNA

16S ribosomal  (rRNA) sequencing using next generation sequencing is a method used to identify and compare bacteria and archaea present within almost any type of sample. 16S rRNA gene sequencing is a well-established method for studying phylogeny and taxonomy of samples from complex microbiomes or environments that are difficult or impossible to study.





16s sequencing illumina or PGM low cost prices with MR DNA

MR DNA is a next generation sequencing provider with low cost 16s sequencing services.


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