From Sequence to Knowledge (Phage Genomics Workshop Intro at the 22nd Biennial Evergreen Phage Meeting)

From Sequence to Knowledge
Assembly, Annotation, and Analysis
of Phage genomes from Genomic
and Metagenomic Data Sets
A helping hand through
The Annotation Bottleneck
Ramy K. Aziz

Workshop presenters
6 Aug 2017 Phage Genomics - Evergreen 2017
Alejandro Reyes
AR
Ramy Aziz
RA
Jason Gill
JG

PRELUDE

A bit of history…
• Since 2009, the Genomics Workshop has
become an essential part of the Evergreen
phage meeting
• The challenge always is: how to meet
needs/expectations that are so many and
so diverse, in ~4 hours

A bit of history…
• Since 2009, the Genomics Workshop has
become an essential part of the Evergreen
phage meeting
• The challenge always is: how to meet
needs/expectations that are so many and
so diverse, in ~4 hours
• The answer is:
…….

A bit of history…

The 2011 workshop

The 2013 workshop

The 2015 workshop

MOTIVATION

“The analysis bottleneck”
• Observation:
– We generate more data than we can analyze.
– We generate sequence data faster than
we can analyze them.
• Opinion:
– Not all bottlenecks are
created equal!
– It is important to define the question(s)
before working on the answer(s)!

• The Lavigne paradox (2013)

• The Lavigne paradox (2015)

AUDIENCE

Workshop audience
• Who (how many) among you have:
– annotated at least a phage genome?
– worked on a viral metagenome?
– used the command line (Unix, Linux, Mac
Terminal) for sequence analysis?
• We have actually ran an online survey,
and here is what we found …

Workshop audience

Quick group activity
Defining the question(s):
• Introduce yourself, your institution, and your
favorite phage
• Do you have a genome sequenced? Planning to?
– Why have you sequenced your phage genome?
– Why you want to sequence your phage genome?
• What is the single most pressing question you
want to have answered from genome analysis?

DEFINING THE QUESTION(S)

What you want …... is
from genome from metagenome
Incomplete
frameshift
- complete
- accurate
Credit: Andrew Kropinski Credit: Bas Dutilh
faulty assembly

What you want …... is
from genome from metagenome
6 Aug 2017
Incomplete faulty assembly
frameshift
- complete
- accurate
Phage Genomics - Evergreen 2017
Credit: Andrew Kropinski Credit: Bas Dutilh

A process of reconstruction

• Experimentally
DNA
TGATTGTGTGTTTGCGCAATGCG
ATGTGTATATATAGTGAGCTTGCCC
GTCTCTCTNNNTCTCTTG
TGATTGGTCTNNNTCTCTTGCGCAATGCG

• Experimentally
• Computationally
GTCTCTCTNNNTCTCTTG
DNA
GTCTCTCTNNNTCTCTTG

• Experimentally
• Computationally
GTCTCTCTNNNTCTCTTG
“Any phage
one can get!”
“eDNA”
GTCTCTCTNNNTCTCTTG

Assembly
Gene finding/
ORF calling
tRNA calling
Annotation
(Assigning
functions)
orienting
Validation
Fixing frameshifts
Introns and Inteins Subsystem
assignment
Refinement/
Secondary
annotation
loop
Special purpose:
toxins, morons, integrases,
lifestyle prediction
Regulatory elements
(promoters, terminators)
Output: files and graphics
From Sequence to Knowledge
From raw sequence data to
genome submission/ publication

Classification
• The phage sequence space (Lima-Mendez et al.)
• The phage proteomic tree (Edwards & Rohwer)
• New: VIP tree http://www.genome.jp/viptree

Countless tools

This workshop: outline
1. Annotation overview
2. Automated tools for genome annotation:
– PhAnToMe/RAST related tools
– Galaxy/ Apollo
3. Tools for metagenome-based analyses
– Assembly
– Functional prediction via protein families

Where to go from here?
• Part I:
General introduction of genome annotation
• Part II:
Two levels
– Level 1: Novices and beginners:
Automated annotation tools
– Level 2: Intermediate to advanced users:
Command-line based tools

Online resources/ Slideshare
• Data & links:
– http://egybio.net/tutorial
• Slides
– http://bit.ly/annotation2016
– http://bit.ly/phantome4
– Old tutorials (more detailed, but missing latest ):
• Evergreen 2011: http://slidesha.re/phantome1
• http://slidesha.re/phiRAST1 (by Karin Holmfeldt)
• Evergreen 2013: http://bit.ly/phantome2
• Evergreen 2015: http://bit.ly/phantome3

From Sequence to Knowledge (Phage Genomics Workshop Intro at the 22nd Biennial Evergreen Phage Meeting)

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