FISH510: Applications of New Sequencing Technologies in Aquatic Sciences

This course will provide students (both field and lab-centric) a forum to discuss innovations in sequencing technologies that have recently evolved to play major roles in physiology, ecology, and population genetics.  The format will include presentations from researchers using the technologies, discussions of challenges unique to working with non-model organisms, and a survey of tools available for data analysis.

Today

• Evolution of the class

• Personal Learning Environment

• Sharing what you know

• Biology

• What is new sequencing technology?

Evolution of this class

me

Evolution

Evolution

Evolution

Evolution

Evolution

This class

• Focus on application of large-scale sequencing data to address issues in ecology and natural resource management

Large scale sequence data

•Large

•Technologies are constantly changing*

•Bioinformatics is not trivial

Today

• Evolution of the class

• Personal Learning Environment

• Sharing what you know

• Biology

• What is new sequencing technology?

Large scale sequence data

•Large

•Technologies are constantly changing*

•Bioinformatics is not trivial

Teaching you to teach yourself

P L E

P L E

P L E

P L E

me

Networks

Directionality

• Push

• Pull (~passive)

• Pull (active) eg ask question

Where would you pull from?

Databases

other aquaticy databases

other aquaticy databases

what (where) else ?

how could you push?

how could you push without and effort?

Evolution

Today

• Evolution of the class

• Personal Learning Environment

• Sharing what you know

• Biology

• What is new sequencing technology?

Evolution

C L E

Today

• Evolution of the class

• Personal Learning Environment

• Sharing what you know

• Biology

• What is new sequencing technology?

Why do nucleotides even matter?

BiologyGenetics

Are these populations reproductively isolated or is there successfully interbreeding?

Transcriptomics

Has temperature stress compromised immune function?

What makes a parasite virulent?

How are growth processes controlled?

Biology

Gene Expression is a Dynamic Process

•Different proteins perform a wide variety of biological functions.

• enzymes, which catalyze chemical reactions • structural or mechanical roles, • immune response • storage and transport of chemical signals• cell communication

Genes are the expressed portion of the genome

Genomic DNA

Messenger RNA (mRNA)

mRNA codes for proteins

BiologyWhat can influence gene expression…

Environment (Temperature, Salinity, Oxygen)Pollution (Environmental Estrogens, Pesticides)Development and Everyday Biology

Gene Expression is a Dynamic Process

•Different proteins perform a wide variety of biological functions.

• enzymes, which catalyze chemical reactions • structural or mechanical roles, • immune response • storage and transport of chemical signals• cell communication

Genes are the expressed portion of the genome

Genomic DNA

Messenger RNA (mRNA)

mRNA codes for proteins

BiologyWhat can influence gene expression…

Environment (Temperature, Salinity, Oxygen)Pollution (Environmental Estrogens, Pesticides)Development and Everyday Biology

Gene Expression is a Dynamic Process

•Different proteins perform a wide variety of biological functions.

• enzymes, which catalyze chemical reactions • structural or mechanical roles, • immune response • storage and transport of chemical signals• cell communication

Genes are the expressed portion of the genome

Genomic DNA

Messenger RNA (mRNA)

mRNA codes for proteins

EVERYTHING

Biology

link

Biology

Biology

BiologyStructure of DNA

Purines Pyrimidines

BiologySense Strand

Antisense Strand

Biology

Transcription changes over timeRNA is less stable than DNA

RNA IsolationProcess immediatelyPreserve itDisrupt sample to release RNA Extract and purify acid phenol/choloroform glass filter oligo dT bindingStore

1. Reverse Transcribe RNA into complementary DNA

2.

This allows PCR to be performed on RNA

Working with RNA

Today

• Evolution of the class

• Personal Learning Environment

• Sharing what you know

• Biology

• What is new sequencing technology?

NGS

NGS

NGS

• template prep

• sequencing and imaging

• data analysis

template prep

• clonal amplification of single DNA molecules

• single DNA molecule template

clonally amplified

• emulsion PCR

• solid-phase amplification

emulsion PCR

solid phase

single molecule

Sequencing and imaging

• Cyclic reversible termination

• Sequencing by ligation

• Pyrosequencing

CRT

CRT

Sequencing and imaging

• Cyclic reversible termination

• Sequencing by ligation

• Pyrosequencing

SBL

Sequencing and imaging

• Cyclic reversible termination

• Sequencing by ligation

• Pyrosequencing

pyrosequencing

Genome Enrichment What?Why?

some ngs applications•variant discovery by resequencing targeted regions of interest or whole genomes,• de novo assemblies of bacterial and lower eukaryotic genomes, •cataloguing the transcriptomes of cells, •tissues and organisms (RNA–seq)5, •genome-wide profiling of epigenetic marks and chromatin structure using other seq-based methods (ChIP–seq, methyl–seq and DNase–seq)4,•species classification and/or gene discovery by metagenomics studies7

videos

You should know

• know what this class is going to be

• how you are going to participate

• what you can learn