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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.





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37. PLoS One. 2014 Sep 24;9(9):e106491. doi: 10.1371/journal.pone.0106491.

eCollection 2014.


Diversity and community composition of methanogenic archaea in the rumen of

Scottish upland sheep assessed by different methods.


Snelling TJ(1), Genç B(2), McKain N(1), Watson M(3), Waters SM(4), Creevey CJ(5),

Wallace RJ(1).


Author information:

(1)Rowett Institute of Nutrition and Health, University of Aberdeen, Bucksburn,

Aberdeen, United Kingdom. (2)Department of Animal Nutrition and Nutritional

Diseases, Faculty of Veterinary Medicine, Ondokuz Mayis University, Samsun,

Turkey. (3)ARK Genomics, The Roslin Institute, Easter Bush, Midlothian, United

Kingdom. (4)Animal and Bioscience Research Department, Animal and Grassland

Research and Innovation Centre, Teagasc, Grange, Dunsany, Co. Meath, Ireland.

(5)Institute of Biological, Environmental and Rural Sciences, Aberystwyth

University, Aberystwyth, Ceredigion, United Kingdom.


Ruminal archaeomes of two mature sheep grazing in the Scottish uplands were

analysed by different sequencing and analysis methods in order to compare the

apparent archaeal communities. All methods revealed that the majority of

methanogens belonged to the Methanobacteriales order containing the

Methanobrevibacter, Methanosphaera and Methanobacteria genera. Sanger sequenced

1.3 kb 16S rRNA gene amplicons identified the main species of Methanobrevibacter

present to be a SGMT Clade member Mbb. millerae (≥ 91% of OTUs); Methanosphaera

comprised the remainder of the OTUs. The primers did not amplify ruminal

Thermoplasmatales-related 16S rRNA genes. Illumina sequenced V6-V8 16S rRNA gene

amplicons identified similar Methanobrevibacter spp. and Methanosphaera clades

and also identified the Thermoplasmatales-related order as 13% of total archaea.

Unusually, both methods concluded that Mbb. ruminantium and relatives from the

same clade (RO) were almost absent. Sequences mapping to rumen 16S rRNA and mcrA

gene references were extracted from Illumina metagenome data. Mapping of the

metagenome data to 16S rRNA gene references produced taxonomic identification to

Order level including 2-3% Thermoplasmatales, but was unable to discriminate to

species level. Mapping of the metagenome data to mcrA gene references resolved

69% to unclassified Methanobacteriales. Only 30% of sequences were assigned to

species level clades: of the sequences assigned to Methanobrevibacter, most

mapped to SGMT (16%) and RO (10%) clades. The Sanger 16S amplicon and Illumina

metagenome mcrA analyses showed similar species richness (Chao1 Index 19-35),

while Illumina metagenome and amplicon 16S rRNA analysis gave lower richness

estimates (10-18). The values of the Shannon Index were low in all methods,

indicating low richness and uneven species distribution. Thus, although much

information may be extracted from the other methods, Illumina amplicon sequencing

of the V6-V8 16S rRNA gene would be the method of choice for studying rumen

archaeal communities.


DOI: 10.1371/journal.pone.0106491

PMCID: PMC4175461

PMID: 25250654  [PubMed - indexed for MEDLINE]



38. PLoS One. 2014 Sep 10;9(9):e106707. doi: 10.1371/journal.pone.0106707.

eCollection 2014.


A comparative metagenome survey of the fecal microbiota of a breast- and a

plant-fed Asian elephant reveals an unexpectedly high diversity of glycoside

hydrolase family enzymes.


Ilmberger N(1), Güllert S(1), Dannenberg J(1), Rabausch U(1), Torres J(1),

Wemheuer B(2), Alawi M(3), Poehlein A(2), Chow J(1), Turaev D(4), Rattei T(4),

Schmeisser C(1), Salomon J(5), Olsen PB(5), Daniel R(2), Grundhoff A(6), Borchert

MS(5), Streit WR(1).


Author information:

(1)Universität Hamburg, Biozentrum Klein Flottbek, Abteilung für Mikrobiologie &

Biotechnologie, Hamburg, Germany. (2)Georg-August-University Göttingen, Institute

of Microbiology and Genetics, Göttingen, Germany. (3)University Medical Center

Hamburg-Eppendorf, Bioinformatics Service Facility, Hamburg, Germany.

(4)University of Vienna, CUBE - Division for Computational Systems Biology,

Department of Microbiology and Ecosystem Science, Vienna, Austria. (5)Novozymes

A/S, Microbial Discovery, Bagsværd, Denmark. (6)Heinrich Pette Institute, Leibniz

Institute for Experimental Virology, Hamburg, Germany.


A phylogenetic and metagenomic study of elephant feces samples (derived from a

three-weeks-old and a six-years-old Asian elephant) was conducted in order to

describe the microbiota inhabiting this large land-living animal. The microbial

diversity was examined via 16S rRNA gene analysis. We generated more than 44,000

GS-FLX+454 reads for each animal. For the baby elephant, 380 operational

taxonomic units (OTUs) were identified at 97% sequence identity level; in the

six-years-old animal, close to 3,000 OTUs were identified, suggesting high

microbial diversity in the older animal. In both animals most OTUs belonged to

Bacteroidetes and Firmicutes. Additionally, for the baby elephant a high number

of Proteobacteria was detected. A metagenomic sequencing approach using Illumina

technology resulted in the generation of 1.1 Gbp assembled DNA in contigs with a

maximum size of 0.6 Mbp. A KEGG pathway analysis suggested high metabolic

diversity regarding the use of polymers and aromatic and non-aromatic compounds.

In line with the high phylogenetic diversity, a surprising and not previously

described biodiversity of glycoside hydrolase (GH) genes was found. Enzymes of 84

GH families were detected. Polysaccharide utilization loci (PULs), which are

found in Bacteroidetes, were highly abundant in the dataset; some of these

comprised cellulase genes. Furthermore the highest coverage for GH5 and GH9

family enzymes was detected for Bacteroidetes, suggesting that bacteria of this

phylum are mainly responsible for the degradation of cellulose in the Asian

elephant. Altogether, this study delivers insight into the biomass conversion by

one of the largest plant-fed and land-living animals.


DOI: 10.1371/journal.pone.0106707

PMCID: PMC4160196

PMID: 25208077  [PubMed - indexed for MEDLINE]



39. BMC Microbiol. 2014 Sep 7;14:225. doi: 10.1186/s12866-014-0225-3.


The presence of nitrate dramatically changed the predominant microbial community

in perchlorate degrading cultures under saline conditions.


Stepanov VG, Xiao Y, Tran Q, Rojas M, Willson RC, Fofanov Y, Fox GE, Roberts DJ.


BACKGROUND: Perchlorate contamination has been detected in both ground water and

drinking water. An attractive treatment option is the use of ion-exchange to

remove and concentrate perchlorate in brine. Biological treatment can

subsequently remove the perchlorate from the brine. When nitrate is present, it

will also be concentrated in the brine and must also be removed by biological

treatment. The primary objective was to obtain an in-depth characterization of

the microbial populations of two salt-tolerant cultures each of which is capable

of metabolizing perchlorate. The cultures were derived from a single ancestral

culture and have been maintained in the laboratory for more than 10 years. One

culture was fed perchlorate only, while the other was fed both perchlorate and


RESULTS: A metagenomic characterization was performed using Illumina DNA

sequencing technology, and the 16S rDNA of several pure strains isolated from the

mixed cultures were sequenced. In the absence of nitrate, members of the

Rhodobacteraceae constituted the prevailing taxonomic group. Second in abundance

were the Rhodocyclaceae. In the nitrate fed culture, the Rhodobacteraceae are

essentially absent. They are replaced by a major expansion of the Rhodocyclaceae

and the emergence of the Alteromonadaceae as a significant community member. Gene

sequences exhibiting significant homology to known perchlorate and nitrate

reduction enzymes were found in both cultures.

CONCLUSIONS: The structure of the two microbial ecosystems of interest has been

established and some representative strains obtained in pure culture. The results

illustrate that under favorable conditions a group of organisms can readily

dominate an ecosystem and yet be effectively eliminated when their advantage is

lost. Almost all known perchlorate-reducing organisms can also effectively reduce

nitrate. This is certainly not the case for the Rhodobacteraceae that were found

to dominate in the absence of nitrate, but effectively disappeared in its

presence. This study is significant in that it reveals the existence of a novel

group of organisms that play a role in the reduction of perchlorate under saline

conditions. These Rhodobacteraceae especially, as well as other organisms present

in these communities may be a promising source of unique salt-tolerant enzymes

for perchlorate reduction.


DOI: 10.1186/s12866-014-0225-3

PMCID: PMC4174249

PMID: 25194715  [PubMed - indexed for MEDLINE]



40. Mol Ecol. 2014 Sep;23(18):4498-510. doi: 10.1111/mec.12885. Epub 2014 Sep 8.


The Sphagnum microbiome supports bog ecosystem functioning under extreme



Bragina A(1), Oberauner-Wappis L, Zachow C, Halwachs B, Thallinger GG, Müller H,

Berg G.


Author information:

(1)Institute of Environmental Biotechnology, Graz University of Technology,

Petersgasse 12, 8010, Graz, Austria.


Sphagnum-dominated bogs represent a unique yet widely distributed type of

terrestrial ecosystem and strongly contribute to global biosphere functioning.

Sphagnum is colonized by highly diverse microbial communities, but less is known

about their function. We identified a high functional diversity within the

Sphagnum microbiome applying an Illumina-based metagenomic approach followed by

de novo assembly and MG-RAST annotation. An interenvironmental comparison

revealed that the Sphagnum microbiome harbours specific genetic features that

distinguish it significantly from microbiomes of higher plants and peat soils.

The differential traits especially support ecosystem functioning by a symbiotic

lifestyle under poikilohydric and ombrotrophic conditions. To realise a

plasticity-stability balance, we found abundant subsystems responsible to cope

with oxidative and drought stresses, to exchange (mobile) genetic elements, and

genes that encode for resistance to detrimental environmental factors, repair and

self-controlling mechanisms. Multiple microbe-microbe and plant-microbe

interactions were also found to play a crucial role as indicated by diverse genes

necessary for biofilm formation, interaction via quorum sensing and nutrient

exchange. A high proportion of genes involved in nitrogen cycle and recycling of

organic material supported the role of bacteria for nutrient supply. 16S rDNA

analysis indicated a higher structural diversity than that which had been

previously detected using PCR-dependent techniques. Altogether, the diverse

Sphagnum microbiome has the ability to support the life of the host plant and the

entire ecosystem under changing environmental conditions. Beyond this, the moss

microbiome presents a promising bio-resource for environmental biotechnology -

with respect to novel enzymes or stress-protecting bacteria.


© 2014 John Wiley & Sons Ltd.


DOI: 10.1111/mec.12885

PMID: 25113243  [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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