Unlike the warm climates

that rule Asian countries,

North America is at the

mercy of harsh climates,

and frost. Productivity, and

yield is as such limited not

only by the crop's

production capacity, but

also temporal factors. In

particular, major crops

such as potatoes – freezing

sensitive – are cultivated in

cold climates which

substantially affects yield.

Between impossibility, and

actuality lies the potential

to genetically alter crops to

serve a defined function.

Where traditional breeding

has failed, biotechnology

has offered much potential

in the generation of crops

that are freezing tolerant.

Typically, this is realized

by conferring a gene -

AtCBF - from an organism

to a selected crop species.

AtCBF1-3, the CBF genes

found in Arabidopsis

thaliana, regulate the

activation of a myriad of

genes responsible for

freezing tolerance. In

particular, CBF genes

(AtCBF1, AtCBF2, and

AtCBF3), a transcription

factor found in Arabidopsis

thaliana, ciphers proteins

that bestows freezing

tolerance upon the plant

via induction of COR (cold

responsive genes).

Constitutive expression of

AtCBF was however found

to be associated with

adverse side effects.

Introduction of a transgene

is at times insufficient for

both resistance, and

productivity. The first

potential solution is to

inspect alternative

transgenes; the second

solution requires

modifications to the

existing system. In this

issue, Pino et al., (2007)

replaced a constitutive

promoter by a stress

inducible promoter to

enable production of the

transgene only under

stressful conditions. By

substituting a constitutive

CaMV 35S promoter with

a stress inducible rd29A

promoter, the authors

manage to bestow upon

Solanum tuberosum both

resistance, and

productivity.

It has been noted that

constitutive expression of

AtCBF – that confer

freezing resistance – leads

to retarded growth,

lowered biomass/foliar

mass, late flowering, and

abolished tuber formation.

The simplest solution to

this problem would be to

substitute the current

transgene with another.

However, it is quite

probable that the novel

transgene will result in the

same problem. In fact,

Kasuga et al. (1999)

utilized the DRE-binding

protein DREB1A, and the

CaMV 35S constitutive

promoter to confer

resistance to freezing,

drought, and salt stresses.

However, constitutive

expression of the transgene

resulted in growth

retardation as well. The

alternative is to modify the

existing system; in this

case, the latter is done by

inducing the transgene

only when required or

under cold stress. In fact, it

has been shown multiple

times that a transgene

associated with a stress

inducible promoter

compared to a constitutive

promoter possess fewer

negative traits such as low

yield, or biomass.

Replacing a constitutive

promoter with a stress-

inducible promoter results

in transgenic Solanum

tuberosum plants that are

both highly productive,

and resistant.

The experiments

conducted by Pino et al.,

(2007) are akin to those

conducted by Kasuga et al.

(1999). Kasuga et al.

(1999) attempted to

compare the phenotype of

the 35S:DREB1A, and the

rd29A:DREB1A lines.

They showed that the

35S:DREB1A line had

fewer seeds, and showed

stunted growth. The

rd29A:DREB1A lines had

mild growth retardation.

Further, Kasuga et al.

(1999) showed that 96.2%

of the rd29A:DREB1A

lines, and 77.9% of the

35S:DREB1A lines

survived after exposure to

cold temperatures. Oddly

enough, the

rd29A:DREB1A lines

outperformed the

35S:DREB1A lines for

many a stress (drought

stress:

35S:DREB1A=39.7-69.2%

survival,

rd29A:DREB1A=76.7%

survival; salinity:

35S:DREB1A=29.4%

survival,

rd29A:DREB1A=78.6%

survival).

Much like Kasuga et al.

(1999), Pino et al., (2007)

altered the genomic unit by

replacing the constitutive

CaMV 35S promoter with

the rd29A stress inducible

promoter. Constitutive

over-expression of AtCBF

is responsible for the side

effects; use of a stress

inducible promoter such as

rd29A can reduce adverse

side effects. Following

substitution of the

constitutive CaMV 35S

promoter for the stress

inducible rd29A promoter,

and transformation using

agrobacterium into S.

tuberosum cv. Umatilla,

explants – derived from

transformed plants – were

propagated in vitro. Tissue

culture was utilized to

regenerate the callus.

Relative to the 35S:AtCBF

lines (constitutive

promoter), the

prd29A:AtCBF lines

showed higher plant, and

tuber mass. Foliar biomass

in the prd29A:AtCBF lines

were unaffected by the

construct.

The paper by Pino et al.,

(2007) implies that

transgenic systems can be

further altered to obtain

desired features instead of

resorting to novel

transgenes. This also

implies that transgenes

alone do not control the

system, but rather that by

modifying a subsection of

a unit (that controls

transcription/translation),

one can tweak the plant’s

genome to perform in a

particular manner. In other

words, this also implies

that when generating

transgenic plants,

introduction of a novel

gene or alteration of

existing genes is one way

of modifying the system;

however, modifications

can be carried out on the

promoter, and the polyA

signal amongst others. In

this paper, the authors

attempt to modify the

promoter in addition to the

transgene, however, it is

quite possible that with

further modifications, this

system can be tweaked.

For instance, it is quite

well known that there

exists a negative

relationship between

resistance, and growth. The

Resource allocation theory

states that due to a

limitation in the resources

available, the plant must

partition said resource

between resistance, or

growth. In such cases, the

activation of a particular

gene under a particular set

of circumstances would

enable the plant to bypass

this limitation. If such is

the case, then one might

add that reducing energy

expended for production of

proteins might aid the

plant. In other words, the

turn-over rate, the efficacy

of the transgene, and the

time of production would

affect the phenotype of the

plant.

References

Kasuga, M., Liu, Q.,

Miura, S., Yamaguchi-

Shinozaki, K., and

Shinozaki, K. (1999).

Improving plant drought,

salt, and freezing tolerance

by gene transfer of a single

stress-inducible

transcription factor. Nature

Biotechnology 17, 287-

291.

Pino, M., Skinner, J., Park,

E., Jeknić, Z., Hayes, P.,

Thomashow, M., and

Chen, T. (2007). Use of a

stress inducible promoter

to drive ectopic AtCBF

expression improves potato

freezing tolerance while

minimizing negative

effects on tuber yield. Plant

Biotechnology J 5, 591-

604.