This document describes a new DNA metabarcoding approach for high-throughput biodiversity assessment using environmental DNA. Short DNA barcode markers are amplified from bulk soil or water samples and sequenced on next-generation platforms to identify all taxa present, without needing to isolate individual organisms. The method was tested on soil samples from an alpine meadow, identifying 8 earthworm species from sequence reads of short mtDNA markers. Future applications could include capturing DNA with hundreds of probes and shotgun sequencing of soil eDNA for comprehensive biodiversity surveys at large scales.
DNA-based methods for bioaerosol analysisjordanpeccia
Information for producing phylogenetic/taxonomic libraries of airborne bacteria and fungi. Includes fundamental background information, approaches for sequencing and data analysis, two case studies, and a review of sampling methods
DNA barcoding is a standardized approach to identifying plants and animals by minimal sequences of DNA, called DNA barcodes.
DNA barcode - short gene sequences taken from a standardized portion of the genome that is used to identify species
and this presentation gives much introducing about DNA barcodes developed for Prokaryotes and Eukaryotes.
Various barcoding genes which are evolutionary conserved.
techniques to develop a DNA bar-code and its future perspectives
Current technologies and future technologies of DNA barcoding. Applications regarding environment awareness. it also contains 2-3 case studies
DNA sequence analysis of a uniform target gene like the mitochondrial cytochrome oxidase subunit I (COI) to enable species identification has been referred to as “DNA Barcoding”, by analogy with the Universal Product Code (UPC) system barcodes used to identify manufactured goods.
DNA barcoding has the potential to be a practical method for identification of the estimated 10 million species of eukaryotic life on earth.
DNA-based methods for bioaerosol analysisjordanpeccia
Information for producing phylogenetic/taxonomic libraries of airborne bacteria and fungi. Includes fundamental background information, approaches for sequencing and data analysis, two case studies, and a review of sampling methods
DNA barcoding is a standardized approach to identifying plants and animals by minimal sequences of DNA, called DNA barcodes.
DNA barcode - short gene sequences taken from a standardized portion of the genome that is used to identify species
and this presentation gives much introducing about DNA barcodes developed for Prokaryotes and Eukaryotes.
Various barcoding genes which are evolutionary conserved.
techniques to develop a DNA bar-code and its future perspectives
Current technologies and future technologies of DNA barcoding. Applications regarding environment awareness. it also contains 2-3 case studies
DNA sequence analysis of a uniform target gene like the mitochondrial cytochrome oxidase subunit I (COI) to enable species identification has been referred to as “DNA Barcoding”, by analogy with the Universal Product Code (UPC) system barcodes used to identify manufactured goods.
DNA barcoding has the potential to be a practical method for identification of the estimated 10 million species of eukaryotic life on earth.
olymerase chain reaction (PCR) is a method widely used in molecular biology to make several copies of a specific DNA segment. Using PCR, copies of DNA sequences are exponentially amplified to generate thousands to millions of more copies of that particular DNA segment.
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PLEASE DOWNLOAD TO SEE ANIMATION.
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Speeding up sequencing: Sequencing in an hour enables sample to answer in a w...Thermo Fisher Scientific
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Ion Torrent semiconductor-based sequencing instruments utilities flow sequencing with speed largely dependent on and the number of nucleotide flows (one flow produces ~0.5 base) and the speed of the flows (Figure 2).
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As an in-depth case study I explain the methods and significance of our 2013 Nature paper on adaptive genotypic molecular convergence in echolocating mammals.
I then highlight some of the avenues of study on the frontiers of current research.
olymerase chain reaction (PCR) is a method widely used in molecular biology to make several copies of a specific DNA segment. Using PCR, copies of DNA sequences are exponentially amplified to generate thousands to millions of more copies of that particular DNA segment.
In Vitro Analog of the Primitive Streak (ANIMATED)Nikolay Turovets
PLEASE DOWNLOAD TO SEE ANIMATION.
In vitro analog of the primitive streak: efficient derivation of highly enriched populations of hepatocytes from various types of human pluripotent stem cells.
May, 2011
How to Standardise and Assemble Raw Data into Sequences: What Does it Mean fo...Joseph Hughes
11th OIE Seminar at the XVII INTERNATIONAL SYMPOSIUM OF THE WORLD ASSOCIATION OF VETERINARY LABORATORY DIAGNOSTICIANS (WAVLD)
Saskatoon - 17th June 2015
National Agricultural Innovation Project (NAIP), ICAR and the International Food Policy Research Institute (IFPRI) organized a two day workshop on ‘Impact of capacity building programs under NAIP’ on June 6-7, 2014 at AP Shinde Auditorium, NASC Complex, Pusa, New Delhi. The main purpose of the workshop was to present and discuss the findings of the impact evaluation study on capacity building programs under NAIP by IFPRI. The scientists from ICAR and agricultural universities were sent abroad to receive training in specialized research techniques. Post-training, scientists were expected to work on collaborative projects within the ICAR, which would further enrich their knowledge and skills, expand their research network and stimulate them’ to improve their productivity, creativity and quality of their research. The ICAR commissioned with IFPRI (International Food Policy Research Institute) to undertake an evaluation of these capacity building programs under NAIP in July 2012. The workshop shared the findings on the impact of capacity building programs under NAIP and evolve strategies for future capacity building programs
Speeding up sequencing: Sequencing in an hour enables sample to answer in a w...Thermo Fisher Scientific
At this time next generation sequencing (NGS) is hindered by slow and often manual workflow procedures. Decreasing overall workflow times is critical for the widespread adoption of targeted and whole genome sequencing (WGS) for many time-sensitive applications, in particular for infectious disease analysis. To this end, we describe improvements to the four main steps of the NGS workflow: i) library preparation; ii) template preparation, iii) sequencing; iv) and data analysis. Together, these advances dramatically decrease the overall turnaround times.
Ion Torrent semiconductor-based sequencing instruments utilities flow sequencing with speed largely dependent on and the number of nucleotide flows (one flow produces ~0.5 base) and the speed of the flows (Figure 2).
Evaluation of Pool-Seq as a cost-effective alternative to GWASAmin Mohamed
Whole-genome sequencing (WGS) of pools of individuals (Pool-Seq) provides a cost-effective method for genome-wide association studies (GWAS), and offers an alternative to sequencing of individuals that remains cost prohibitive. Pool-Seq is being increasingly used in population genomic studies in both model and non-model organisms. In this paper, the ability of Pool-Seq to recover known GWAS signals was evaluated. Existing GWAS data for 2,112 animals with 729K SNPs were obtained and pooled to simulate data obtained from a pooled WGS approach. Traditional GWAS results was compared with the absolute allele frequency difference (dAF) metric suitable for use with Pool-Seq data. Specifically, we tested the ability of dAF scans to recover known GWAS signals for two different traits with large and moderate gene effects. Pools of different sizes (50, 100 and 200 individuals per pool) were also compared. The results showed the ability of the absolute allele frequency difference (dAF) approach to recover known GWAS peaks obtained by traditional SNP association and recommended the use of a pool size of 100 individuals for DNA pooling.
Phylogenomic methods for comparative evolutionary biology - University Colleg...Joe Parker
Invited research seminar given to MSc students at University College Dublin on 24th October 2013.
I introduce the discipline of phylogenomics - comparative phylogenetic analyses of DNA sequences across genomes - and some of the applications and recent breakthroughs in the field.
As an in-depth case study I explain the methods and significance of our 2013 Nature paper on adaptive genotypic molecular convergence in echolocating mammals.
I then highlight some of the avenues of study on the frontiers of current research.
DNA barcoding is a technique in which species identification is performed by using DNA sequences from a small fragment of the genome, with the aim of contributing to a wide range of ecological and conservation studies in which traditional taxonomic identification is not practical.
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A microarray is a laboratory tool used to detect the expression of thousands of genes at the same time. DNA microarrays are microscope slides that are printed with thousands of tiny spots in defined positions, with each spot containing a known DNA sequence or gene.
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'Genomics' is nothing but the study of entire genetic compliment of an organism. Plant genomics is study of plant genome. This is my topic of M.Sc. course 'Plant biotechnology'.
whole genome analysis
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This presentation includes basic of PCOS their pathology and treatment and also Ayurveda correlation of PCOS and Ayurvedic line of treatment mentioned in classics.
June 3, 2024 Anti-Semitism Letter Sent to MIT President Kornbluth and MIT Cor...Levi Shapiro
Letter from the Congress of the United States regarding Anti-Semitism sent June 3rd to MIT President Sally Kornbluth, MIT Corp Chair, Mark Gorenberg
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The US House of Representatives is deeply concerned by ongoing and pervasive acts of antisemitic
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This Congress will not stand idly by and allow an environment hostile to Jewish students to persist. The House believes that your institution is in violation of Title VI of the Civil Rights Act, and the inability or
unwillingness to rectify this violation through action requires accountability.
Postsecondary education is a unique opportunity for students to learn and have their ideas and beliefs challenged. However, universities receiving hundreds of millions of federal funds annually have denied
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The House of Representatives will not countenance the use of federal funds to indoctrinate students into hateful, antisemitic, anti-American supporters of terrorism. Investigations into campus antisemitism by the Committee on Education and the Workforce and the Committee on Ways and Means have been expanded into a Congress-wide probe across all relevant jurisdictions to address this national crisis. The undersigned Committees will conduct oversight into the use of federal funds at MIT and its learning environment under authorities granted to each Committee.
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RPMS TEMPLATE FOR SCHOOL YEAR 2023-2024 FOR TEACHER 1 TO TEACHER 3
Pierre Taberlet - Saturday Closing Plenary
1. The DNA metabarcoding approach
for analyzing environmental
samples: high throughput plant and
animal identification
Pierre Taberlet, Eric Coissac, François Pompanon,
Johan Pansu, Wasim Shehzad, Tiayyba Riaz
Laboratoire d'Ecologie Alpine, CNRS UMR 5553
Université Joseph Fourier, Grenoble, France
2. Need for high throughput
collection of biodiversity data
• For research
• For management
At the moment, no
possibilty to use satellites
for identifying taxa and
collecting biodiversity data.
Why not using DNA
metabarcoding?
NASA Earth Observing System: Terra Satellite Platform
3. Our goal: a new high throughput
approach for obtaining
biodiversity data
(based on the DNA-barcoding concept, and
using next generation sequencers)
• A single sampling in the field
• Simple and robust metabarcoding
experiments at the bench
• Complete biodiversity assessment at
the sampling site
4. Environmental DNA (eDNA)
• Environmental DNA refers to DNA that can be
extracted from air, water, or soil, without isolating
any specific type of organism beforehand
• Two types:
– intracellular eDNA
– extracellular eDNA
• Intracellular eDNA commonly used by
microbiologists
• We focus on extracellular eDNA
5. Constraints of working with
environmental DNA
• Complex mixture containing degraded DNA
• The eDNA extract must be representative of the local
biodiversity
• The standard DNA barcodes are not optimal (they are by
far too long to reveal the whole spectrum of biodiversity)
• The primers must be highly versatile (to equally amplify
the different target DNAs)
• Problem of the taxonomic resolution when using very
short barcodes
• At the moment, problem of the reference database when
using non-standard barcodes
6.
7. "Roche Noire" experiment
in French Alps
• Four plant communities
– dry high alpine meadows dominated by Kobresia myosuroides
– low alpine meadows dominated by Carex sempervirens
– subalpine heath dominated by Vaccinium sp.
– subalpine grasslands dominated by Festuca paniculata
• Three plots per plant community (12 plots)
• Two soil samples per plot (with 80 cores per sample)
• Two DNA extractions per sample
• Two DNA amplifications per extraction
8. "Roche Noire" experiment in
French Alps
80 soil cores per sample
10m
● Extraction of extracellular DNA from kilograms of soil using a phosphate buffer
● DNA amplification of the P6 loop of the chloroplast trnL (UAA) intron
● Sequencing on the 454
9.
10.
11.
12.
13. "Roche Noire" experiment: projections
of a between class analysis
Axe 2 (15.4%) Axe 3 (13.2%)
Axe 1 (18.9%) Axe 2 (15.4%)
Carex
Festuca
Kobresia
Vaccinium
A B
Taberlet P, Prud'homme S, Campione E, et al. (2012) Extraction of extracellular DNA from large
amount of soil for metabarcoding studies. Molecular Ecology, 21, in press.
doi: 10.1111/j.1365-1294X.2011.05317.x.
14. Simple and robust
metabarcoding experiments
• In silico analysis: design and test of short
metabarcodes (ecoPrimers, ecoPCR)
• Empirical experiments
– DNA amplification with barcode primers
– Sequencing of the PCR products on next generation
sequencers
• Sequence analysis
– OBITools (www.prabi.grenoble.fr/trac/OBITools)
Ficetola GF, Coissac E, Zundel S, et al. (2010) An in silico approach for the evaluation of DNA
barcodes. BMC Genomics, 11, 434.
Riaz T, Shehzad W, Viari A, Pompanon F, Taberlet P, Coissac E (2011) ecoPrimers: inference of
new DNA barcode markers from whole genome sequence analysis. Nucleic Acids Research,
doi:10.1093/nar/gkr1732.
15. A collection of metabarcoding primers
Taxonomic group Gene Length Accuracy (Bs)
Angiosperms/Gymnosperms cpDNA trnL intron 10-100 bp Genus/Species
Poaceae ITS1 54-88 bp Species
Fungi ITS1 ~ 200 bp Species ?
Vertebrates mtDNA 12S V05 76-110 bp Genus/Species
Teleost fishes mtDNA 12S 60-70 bp Species
Batrachia mtDNA 12S ~ 42-57 bp Species
Earthworms mtDNA 16S (ewB/ewC) ~ 30 bp Species
Earthworms mtDNA 16S (ewD/ewE) ~ 70 bp Species
Oligochaetes mtDNA 16S (ewB/ewE) ~ 120 bp Species
Arthropods/Mollusks mtDNA 16S 35-40 bp Family/Genus
Termites mtDNA 12S ~ 30 bp Species ?
Termites mtDNA 12S ~ 70 bp Species ?
Collembola mtDNA 12S 39-44 bp Species ?
Collembola mtDNA 12S 125-138 bp Species ?
More information soon on www.metabarcoding.org
16. Earthworms from soil DNA
• Eight soil samples collected per plot
• Universal short metabarcodes for earthworms
• Reference database built using samples identified with the
standardized COI barcoding approach
• Sequencing on Illumina GA IIx
d e
b c
mtDNA 12S
30 bp 70 bp
18. Current limitations of the PCR-
based approach
• Dependency on PCR
– Amplification introduces errors
– Difficulty to find suitable barcodes
– Different groups of organisms are analyzed
separately
• Lack of comprehensive taxonomic reference
databases for non-standard metabarcodes
• Limitations linked to the use of organellar
markers
19. Future: capture
• Easier to find a single
conserved region for designing
the probe for the capture than
two close conserved regions for
PCR
• ecoProbes: computer program
for designing suitable probes
(comparable to ecoPrimers)
• Possibility to use hundreds of
probes at the same time
• Both organellar and nuclear
DNA can be analyzed at the
same time
e.g. Briggs AW, Good JM, Green RE, et al. (2009) Targeted retrieval and analysis of five Neandertal
mtDNA genomes. Science, 325, 318-321.
20. An idea of the HiSeq 2000
production per run
• 6 billions of reads of 100 bp
• 6 lines per read
• 55 lines per page (time 11)
• 654 545 454 pages
• 194 400 km long
• 70.5 km high
• more than 3,000 tons of
paper
21. Future: shotgun sequencing
• Shotgun sequencing of soil extracellular
DNA on HiSeq 2000
• We do not know the percentage of
informative reads
• Might allow to use the standard barcode
reference libraries
• Real bioinformatics challenge
• Ongoing experiments…
22. Acknowledgements
Rike Bienert, Kari Anne Bråthen, Christian Brochmann, Anne Krag Brysting, Etienne
Campione, Corinne Cruaud; Francesco de Bello, Tony Dejean, Mary Edwards,
Francesco Ficetola, Frédérick Gavory, Ludovic Gielly, James Haile, Christelle Melo de
Lima, Christian Miquel, Stéphanie Pellier-Cuit, Sophie Prud'homme, Delphine Rioux,
Julien Roy, Jorn Henrik Sønstebø, Wilfried Thuiller, Alice Valentini, Eske Willerslev,
Patrick Wincker, Nigel Yoccoz
23. Thank you for
your attention
Contacts: eric.coissac@inria.f; pierre.taberlet@ujf-grenoble.fr
Molecular Ecology will publish in 2012 a special issue on Environmental DNA