Primary Metabolism

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Primary Metabolism
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Microbial life strategies
Nomads
Settlers
1 mm
3 cm
10 cm
Actinomycetes Cyanobacteria Filamentous Fungi
E. coli Salmonella Streptococcus
Settlers have to protect themselves from the
Nomads
Antibiotics
Signaling molecules
Siderophores
Specialized cell walls/lipids
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Chemical Genetics
Secondary metabolite
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The Genus Salinispora
Jensen, Fenical
Udwary, Ziegler, Lapidus, Moore, Jensen
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A Yersiniabactin Cluster?
unknown
transport
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The quick brown fox jumped over the lazy dog
Yoda speak Over the lazy dog the quick brown fox
jumped.
A fast tan dingo leaped above some sleepy mutt
Low sequence identity, Similar product
Yersiniabactin cluster from Yersinia pestis
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The quick brown
fox
jumped over the lazy
dog
High sequence identity, But different product
The quick brown
fox
jumped over the lazy
dog
BLAST cant really tell the difference
Would lead to different, but similar, chemical
products that may have different activity
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Project 1 Operon/Cluster identification tool
Rationale I have a need to locate conserved or
related operons or gene clusters across multiple
species. Important to understanding of genome
evolution, and functional analysis.
Goal Develop a rapid automated method to
identify conserved gene clusters or operons
across genomes.
Personnel needs Programmer/scripter (Java or
Perl is ideal) Biologist with some knowledge of
gene structure or genomics
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Identifying Gene Clusters
PKS KS AT DH ER KR ACP Methyl transferase
MT TE NRPS C A T (PCP) Epimerization C-cycli
zation N-methyltransferase TE Other Prenyl
transferase Cytochrome P450 Type III PKS KAS
III
DNA sequence(s)
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Genome
KS AT DH ER KR ACP Methyl transferase MT TE C A T
(PCP) Epimerization C-cyclization N-methyltransfer
ase TE Prenyl transferase Cytochrome P450 KAS III
Library of query genes and desirability scores
Score for each gene
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Project 2 Synthetic Sequence Generator
Rationale Synthesis of large DNA sequences is
commercially feasible. There is need for an
organism-agnostic design tool.
Goal Develop a tool to generate
organism-specific DNA sequences suitable for DNA
synthesis from protein sequences
Personnel needs Programmer/scripter Biologist
with some knowledge of molecular biology and
genetics
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Problem I want to understand what the four-gene
operon from each organism does, biochemically.
NEED all four proteins active and in the same
place. Plus, one organism is not available to me.
Old Way
Dans Way
Type a DNA sequence Email sequence to a company
to synthesize it Transform into expression
host Culture organism Evaluate activity in
vivo
PCR-amplify gene(s) Purify PCR product(s) Clone
into expression plasmid(s) Confirm accuracy of
sequence(s) Transform into expression
host Culture organism Purify proteins Evaluate
activity in vitro
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Protein sequence
Codon usage table of host organism
Re-engineered, optimized DNA sequence