Molecular Approaches for Transgene Containment and Their Potential Applications in Horticultural Crops - Transgenic Horticultural Crops: Challenges and Opportunities - page 293

(CRES-T).to.repress.expression.of.four.transcription.factor.genes,.namely. APETALA3 ,.
AGAMOUS ,. LEAFY ,.and. AtMYB26 .that.are.involved.in.the.control.of.loral.mer-
istem. and. organ. identities,. and. the. regulation. of. anther. dehiscence,. respectively..
Transgenic.plants.expressing.each.chimeric.silencer.gene.construct.were.sterile,.and.
resembled.the.loss-of-function. phenotype. of. each. corresponding. gene.. One. advan-
tage.of.using.these.loral.organ.genes.to.create.sterility.is.that.the.products.of.loral.
organ. development. genes. have. no. toxic. effects. on. human. and. animal. health. and.
therefore.there.should.be.few.concerns.if.used.in.edible.horticultural.crops..However,.
ectopic.expression.of.loral.organ.development.genes.frequently.leads.to.alterations.
in.loral.morphology..For.horticultural.crops,.alteration.in.loral.organ.morphology.
can.be.desirable.or.undesirable.depending.on.plant.species.and.applications.
G eneSafe t echnoloGIeS
Genesafe. technologies,. also. called. genetic. use. restriction. technologies. (GURTs),.
involve.the.use.of.genetic.switch.mechanisms.to.control.expression.of.a.toxin.gene.
in. seeds.(see.review.by.Kausch.et.al. 14 )..It.is.an.extension.of.sterility.technologies.
discussed.in.the.“Male.and.female.sterility”.section.of.this.chapter..The.best.example.
of. the. genesafe. technologies. is. the. one. invented. by. Oliver. et. al., 58 . many. called. it.
the. “terminator”. seed. technology.. The. strategy. uses. a. late.embryogenesis. speciic.
gene.promoter.to.control.the.expression.of.a.lethal.gene,.the.bacterial.barnase.gene..
However,.the.barnase.gene.is.interrupted.by.a.short.blocking.DNA.sequence.so.that.
it.is.inactive..The.blocking.DNA.sequence.is.lanked.by.two.loxP.sequences.in.the.
same.orientation..LoxP.is.the.recognition.sequence.of.the.Cre.recombinase.from.the.
bacteriophage. P1. loxP/Cre . DNA. recombination. system. 59,60 . The. interrupted. lethal.
gene.is.introduced.into.one.parental.plant.line..The. Cre .gene.that.is.under.the.control.
of.a.chemically.induced.gene.promoter.such.as.the.tetracycline.inducible.promoter.
is.introduced.into.another.parental.line..Both.of.these.plant.lines.can.produce.viable.
seeds. if. self-pollinated.. If. the. lethal/barnase.line. and.the. Cre . line. are. hybridized,.
their.progeny.should.contain.both.the.chemically.inducible. Cre .gene.and.the.inter-
rupted. barnase. gene. that. is. under. the. control. of. a. late-embryogenesis. active. gene.
promoter..If.hybrid.seeds.produced.from.the.cross.of.these.two.plant.lines.are.treated.
with.the.chemical.inducer,.the.Cre.enzyme.will.be.produced.in.seeds.and.the.interrupt-
ing.DNA.sequence.should.be.deleted..As.a.consequence,.excision.of.the.interrupting.
DNA.sequence.should.make.the.barnase.gene.functional.in.mature.embryos.during.
the.seed.development..If.these.resulting.seeds.are.used.for.planting,.when.the.plants.
derived.reach.the.reproductive.stage,.the.lethal.gene.should.be.expressed.speciically.
in.mature.embryos..The.barnase.gene.expression.in.mature.embryo.will.cause.the.
death.of.embryo.and.thus.prevent.seed.germination.
Similar. to. the. “terminator”. seed. technology,. Kuvshinov. et. al. 61 . described. a.
“recoverable.block.of.function”.(RBF).technology..A.lethal.gene,.also.the.barnase.
gene,.was.driven.by.a.germination-speciic.promoter,.and.the.barstar.(an.inhibitor.
of.barnase).gene.was.placed.under.the.control.of.a.heat-shock.promoter..Under.natural.
conditions,. the. barnase. gene. will. be. expressed. during. seed. germination. but. the.
barstar.gene.will.be.inactive.because.the.heat-shock.promoter.is.not.induced..The.
expression.of.the.barnase.gene.in.germinating.seeds.will.prevent.seed.germination..