GAURAV KUMAR PANDITM.Sc. Plant Biology &

Biotechnology16LPMS 09School of Life Science

University of Hyderabad

gauravbiologist@uohyd.ac.inSupervisor

PROF. C.H.VENKETRAMANA

METAGENOMICS

3rd March 2017

The pure culture

Culture is not enough

Most prokaryotes are extremely difficult to retrieve in a pure culture.

Metagenomics – the key to uncultured microorganisms.

Study of large populations of microoganisms that reproduce clonally from a single cell

 Viromes : Metagenomic sequencing is particularly useful in the study of viral communities.A metagenomic pipeline called Giant Virus Finder showed the first evidence of existence of giant viruses in a saline desert and in Antarctic dry valleys .

Isolate

Metagenomics

CommunityGenomics and Metagenomics

Traditional microbial genomics

Metagenomics “The application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species” - Kevin Chen and Lior Pachter Also referred as Environmental genomics, Ecogenomics, community genomics. The term "metagenomics" was first used by Jo Handelsmann, Jon Clardy, Robert M. Goodman.Reference : Chen K and Pachter L ; Bioinformatics for Whole- Genome Shotgun Sequencing of Microbial Communities, PLoS Comput Biol.2005 Jul;1(2):106-12.

DNA fragmentation (3 Kpb / 30-40 Kpb ) fosmid /cosmid

(35-40 Kpb)

eDNA

plasmid

cloning

Eschericchia coli

transformation

Microtiter plateMetagenomic Libraries

Gene “repository”

Classic Metagenomics

Sequencing primers

Sequencing primers

Small insert vectors

Large insert vectors

eDNA

3Kbp

Next Generation SequencingBiological sample preparation

a.Emulsion PCR b.Bridge PCR

Cyclic Array Sequencing 1.Pyrosequencing (454) 2. SOLiD(by ligation) 3.  Illumina

References:https://www.researchgate.net/figure/268816439_fig1-3,

TWO APPROACHES FOR METAGENOMICS

Functional based screening

Sequence based screening

Reference :www.lucigen.com & metagenomicsrevealed.yolasite.com

LIMITATIONS OF TWO APPROACHES The sequence driven approach

limited existing knowledge: if a metagenomic gene does not look like a gene of known function deposited in the databases, then little can be learned about the gene or its product from sequence alone.

The function driven approach most genes from organisms in wild

communities cannot be expressed easily by a given surrogate host

Therefore, the two approaches are complementary and should be pursued in parallel.

Why do Metagenomics ?Metagenomics may answer Some questions.

How do different species interact?

Can lateral gene transfer be detected?

References : The Metagenomics Group at CBS

UnderstandingMetabolism

Defining the Minimal Gene Set

Genome Engineering

Understanding Cell Structure & Function

Understanding Host Interactions

UnderstandingProtein-Protein

Interactions

UnderstandingExpression

(RNA/Protein)

Discover DNA Variation, Genotyping

Forensics

Drug/VaccineDevelopment

Why do Metagenomics ?

LIMITATIONS Too much data? Most genes are not identifiable Contamination, chimeric clone sequences Requires proteomics or expression studies to

demonstrate phenotypic characteristics Need a standard method for annotating

genomes Requires high throughput instrumentation – not

readily available to most institutions Errors in assembly due to inter-species

similarities.

To identify new enzymes & antibiotics

To assess the effects of age, diet, and pathologic states on the distal gut micro biome of humans living in different environments.

Study antibiotic resistance in uncultured microbes

Improved bioinformatics will quicken analysis for library profiling .

Discoveries such as phylogenic tags (rRNA genes, etc) will give momentum to the growing field.

Learning novel pathways will lead to knowledge about the current nonculturable bacteria to then culture these systems.

FUTURE OF METAGENOMICS

References :www.cev.washington.edu/story/Ecogenomic_Sensor

The illustration was developed in collaboration with Ginger Armbrust and John Delaney, School of Oceanography, University of Washington

Ecogenomic Sensor

Source :John Delaney and CEV. UWash

Video Collected by ROV

Sony Digital BetaCAM Recorder

SDI Over Fiber

SDI Over Fiber Capture

Control

Processing on Cluster

Detection & Classification

Working Flow Diagram of FOCO

SDI = Serial Digital Interface

Conclusion Microorganisms are ubiquitous in nature. Although

we cannot usually see them, microorganisms are essential for every part of life on the Earth. Every process in the biosphere is touched by the microorganisms that convert the key elements of life—carbon, nitrogen, oxygen, and sulfur—into forms accessible to all other living things.

Metagenomics provides a new way of examining the microbial world that has the potential to revolutionize understanding of the entire living world.

Metagenomics combines the power of genomics, bioinformatics, and systems biology.

It provides new access to the microbiological world and uncultured microorganisms.

REFERENCES Thomas T, Gilbert J and Meyer F ;

Metagenomics - a guide from sampling to data analysis ; Microb Inform Exp. 2012 Feb 9;2(1):3.

Chen K and Pachter L ; Bioinformatics for Whole-Genome Shotgun Sequencing of Microbial Communities; PLoS Comput Biol.2005 Jul;1(2):106-12.

Streit WR and Schmitz RA ; Metagenomics – the key to the uncultured microbes ; Curr Opin Microbiol. 2004 Oct;7(5):492-8.

Muth TR and McEntee CM ; Undergraduate Urban Metagenomics Research Module; J Microbiol Biol Educ.2014 May 1;15(1):38-40.

Prescott,Harley and Klein’s Microbiology (Book) ; Seventh Edition ;Chapter 15 Microbial Genomics.

ACKNOWLEDGEMENTMy Supervisor

PROF.C.H.VENKETRAMANAI Would like to take this opportunity to THANKS Prof. C.H.VENKETRAMANA , My Superviror who teach me in the documentation and understanding the METAGENOMICS.

DR.IRFAN A.GHAZIDepartment of Plant Science

(Course coordinator)I extend my heart felt gratitude to DR.IRFAN A.GHAZI for giving us exemplous support during the whole tenure of documentation and preparation.

I , Would like to THANK all of Faculties of Department of Plant science (SLS,UOH) who allowing me to present.

MWow ! She’s Gorgeous

Hi; guys ! I’m Bacilli andThis is my wife Spirilla….

……..Hello ! I’m MicrococcusAnd this is Gallionella.

Listen up my fellow bacteria , we will attack together when we are enough in number. So, how many are you ? Start counting.

A Great big Thank you