Harnessing promoter elements to enhance gene editing in plants: perspectives and advances
Plant Biotechnology Journal,
Год журнала:
2025,
Номер
unknown
Опубликована: Фев. 27, 2025
Summary
Genome‐edited
plants,
endowed
with
climate‐smart
traits,
have
been
promoted
as
tools
for
strengthening
resilience
against
climate
change.
Successful
plant
gene
editing
(GE)
requires
precise
regulation
of
the
GE
machinery,
a
process
controlled
by
promoters,
which
drives
its
transcription
through
interactions
factors
(TFs)
and
RNA
polymerase.
While
constitutive
promoters
are
extensively
used
in
constructs,
their
limitations
highlight
need
alternative
approaches.
This
review
emphasizes
promise
tissue/organ
specific
well
inducible
enable
targeted
spatiotemporal
manner
no
effects
on
other
tissues.
Advances
synthetic
biology
paved
way
creation
offering
refined
control
over
expression
augmenting
potential
GE.
The
integration
these
novel
systems
presents
significant
opportunities
conditional
genome
editing.
Moreover,
advent
bioinformatic
artificial
intelligence
is
revolutionizing
characterization
regulatory
elements,
enhancing
our
understanding
roles
plants.
Thus,
this
provides
insights
into
strategic
use
promoter
to
enhance
precision,
efficiency
specificity
GE,
setting
stage
innovative
crop
improvement
strategies.
Язык: Английский
Versatile plant genome engineering using anti-CRISPR-Cas12a systems
Science China Life Sciences,
Год журнала:
2024,
Номер
unknown
Опубликована: Авг. 15, 2024
Язык: Английский
An efficient CRISPR‐Cas12a‐mediated MicroRNA knockout strategy in plants
Plant Biotechnology Journal,
Год журнала:
2024,
Номер
unknown
Опубликована: Окт. 14, 2024
Summary
In
recent
years,
the
CRISPR‐Cas9
nuclease
has
been
used
to
knock
out
MicroRNA
(miRNA)
genes
in
plants,
greatly
promoting
study
of
miRNA
function.
However,
due
its
propensity
for
generating
small
insertions
and
deletions,
Cas9
is
not
well‐suited
achieving
a
complete
knockout
genes.
By
contrast,
CRISPR‐Cas12a
generates
larger
which
could
significantly
disrupt
secondary
structure
pre‐miRNA
prevent
production
mature
miRNAs.
Through
case
OsMIR390
rice,
we
confirmed
that
Cas12a
more
efficient
tool
than
mutants
gene.
To
further
demonstrate
CRISPR‐Cas12a‐mediated
targeted
nine
OsMIRNA
have
different
spaciotemporal
expression
previously
investigated
via
genetic
approaches.
With
CRISPR‐Cas12a,
up
100%
genome
editing
efficiency
was
observed
at
these
loci.
The
resulting
deletions
suggest
robustly
generated
null
alleles
Transcriptome
profiling
mutants,
as
well
phenotypic
analysis
rice
grains
revealed
function
miRNAs
controlling
gene
regulating
grain
quality
seed
development.
This
established
an
plants.
Язык: Английский
PAM‐relaxed and temperature‐tolerant CRISPR‐Mb3Cas12a single transcript unit systems for efficient singular and multiplexed genome editing in rice, maize, and tomato
Plant Biotechnology Journal,
Год журнала:
2024,
Номер
unknown
Опубликована: Окт. 10, 2024
Class
2
Type
V-A
CRISPR-Cas
(Cas12a)
nucleases
are
powerful
genome
editing
tools,
particularly
effective
in
A/T-rich
genomic
regions,
complementing
the
widely
used
CRISPR-Cas9
plants.
To
enhance
utility
of
Cas12a,
we
investigate
three
Cas12a
orthologs-Mb3Cas12a,
PrCas12a,
and
HkCas12a-in
Protospacer
adjacent
motif
(PAM)
requirements,
efficiencies,
profiles
compared
rice.
Among
these
orthologs,
Mb3Cas12a
exhibits
high
efficiency
at
target
sites
with
a
simpler,
relaxed
TTV
PAM
which
is
less
restrictive
than
canonical
TTTV
LbCas12a
AsCas12a.
optimize
Mb3Cas12a,
develop
an
efficient
single
transcription
unit
(STU)
system
by
refining
linker
between
CRISPR
RNA
(crRNA),
nuclear
localization
signal
(NLS),
direct
repeat
(DR).
This
optimized
enables
precise
rice,
for
fine-tuning
gene
expression
promoter
regions.
Further,
introduced
Arginine
(R)
substitutions
Aspartic
acid
(D)
172,
Asparagine
(N)
573,
Lysine
(K)
579
creating
two
temperature-tolerant
variants:
Mb3Cas12a-R
(D172R)
Mb3Cas12a-RRR
(D172R/N573R/K579R).
These
variants
demonstrate
significantly
improved
lower
temperatures
(22
°C
28
°C)
rice
cells,
showing
best
performance.
We
extend
this
approach
developing
STU
systems
maize
tomato,
achieving
biallelic
mutants
targeting
or
multiple
genes
T
Язык: Английский