An Automated Cell-Free Workflow for Transcription Factor Engineering
Holly M. Ekas,
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Brenda Wang,
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Adam D. Silverman
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et al.
ACS Synthetic Biology,
Journal Year:
2024,
Volume and Issue:
13(10), P. 3389 - 3399
Published: Oct. 7, 2024
The
design
and
optimization
of
metabolic
pathways,
genetic
systems,
engineered
proteins
rely
on
high-throughput
assays
to
streamline
design-build-test-learn
cycles.
However,
assay
development
is
a
time-consuming
laborious
process.
Here,
we
create
generalizable
approach
for
the
tailored
automated
cell-free
gene
expression
(CFE)-based
workflows,
which
offers
distinct
advantages
over
in
vivo
reaction
flexibility,
control,
time
data.
Centered
around
designing
highly
accurate
precise
transfers
Echo
Acoustic
Liquid
Handler,
introduce
pilot
validation
strategies
each
stage
protocol
development.
We
then
demonstrate
efficacy
our
platform
by
engineering
transcription
factor-based
biosensors.
As
model,
rapidly
generate
libraries
127
MerR
134
CadR
factor
variants
3682
unique
CFE
reactions
less
than
48
h
improve
limit
detection,
selectivity,
dynamic
range
mercury
cadmium
detection.
This
was
achieved
assessing
panel
ligand
conditions
sensitivity
(to
0.1,
1,
10
μM
Hg
0,
10,
100
Cd
CadR,
respectively)
selectivity
(against
Ag,
As,
Cd,
Co,
Cu,
Hg,
Ni,
Pb,
Zn).
anticipate
that
Echo-based,
can
be
used
accelerate
multiple
workflows
synthetic
biology.
Language: Английский
The design and engineering of synthetic genomes
Nature Reviews Genetics,
Journal Year:
2024,
Volume and Issue:
unknown
Published: Nov. 6, 2024
Language: Английский
Navigating the challenges of engineering composite specialized metabolite pathways in plants
The Plant Journal,
Journal Year:
2025,
Volume and Issue:
121(6)
Published: March 1, 2025
SUMMARY
Plants
are
a
valuable
source
of
diverse
specialized
metabolites
with
numerous
applications.
However,
these
compounds
often
produced
in
limited
quantities,
particularly
under
unfavorable
ecological
conditions.
To
achieve
sufficient
levels
target
metabolites,
alternative
strategies
such
as
pathway
engineering
heterologous
systems
like
microbes
(e.g.,
bacteria
and
fungi)
or
cell‐free
can
be
employed.
Another
approach
is
plant
engineering,
which
aims
to
either
enhance
the
native
production
original
reconstruct
model
system.
Although
increasing
metabolite
promising
strategy,
plants
exotic
pose
significant
challenges
for
genetic
manipulation.
Effective
requires
comprehensive
prior
knowledge
genes
enzymes
involved,
well
precursor,
intermediate,
branching,
final
metabolites.
Thus,
thorough
elucidation
biosynthetic
closely
linked
successful
metabolic
host
systems.
In
this
review,
we
highlight
recent
advances
engineering.
We
focus
on
efforts
engineer
complex,
multi‐step
pathways
that
require
expression
at
least
eight
transient
three
stable
transformation.
Reports
complex
stably
transformed
remain
relatively
scarce.
discuss
major
hurdles
overcoming
them,
followed
by
an
overview
achievements,
challenges,
solutions
reconstitution
through
Recent
including
computer‐based
predictions
offer
platforms
sustainable
plants.
Language: Английский
An Optimized Version of the Small Synthetic Genome (Mini-Synplastome) for Plastid Metabolic Engineering in Solanum tuberosum (Potato)
ACS Synthetic Biology,
Journal Year:
2024,
Volume and Issue:
unknown
Published: Dec. 10, 2024
Plastids
represent
promising
targets
in
plant
genetic
engineering
for
many
biotech
applications,
ranging
from
their
use
as
bioreactors
the
overproduction
of
valuable
molecules
to
installation
transgenes
improving
traits.
For
over
30
years,
routine
methods
plastid
transformation
have
relied
on
homologous
recombination
integrating
vectors.
However,
nonintegrating
episomal
plasmids
recently
received
more
attention
an
innovative
tool
cells.
One
these
novel
technologies
is
mini-synplastome,
plasmid
with
a
chloroplast-specific
origin
replication
(
Language: Английский