E il naufragare m’è dolce in questo mar
Ovvero come raddoppiare il contenuto in zucchero
Barbabietola da zucchero
Concentrazione di Saccarosio
1802: 5%, 2007: 20%
Resa (kg/ha) 1812: 700
1946: 3500 kg
2005: 13000 kg
With a root yield 40 t/ha at 15.5-18% sugar content,  6 -7 t sugar/ha.
Semente: 1946, 20 kg per Ha, 2005: 1kg (di qualità)
Azoto 1953: 240 kg per Ha, 2005: 80 kg
Canna da Zucchero
At harvest (quindi peso fresco) with best management practices
150-175 ton/ha in sub-tropical zone
150-300 tons/ha in tropical zone (depending on length of growth period)
Per es. in Brasile: 80 t/ha fresh weight (70% di umidità).
Sugar content at harvest is usually between 10 and 12 percent of
the cane fresh weight
http://en.wikipedia.org/wiki/Sugarcane
http://it.wikipedia.org/wiki/Barbabietola
http://en.wikipedia.org/wiki/Ethanol_fuel_in_Brazil
Canna da zucchero
Whilst improvements in yield have continued to accrue, in Australia there has
been no increase in sugar content over the last 40 years (Jackson 2005).
1 ettaro = 2.5 acri 1 ac = 0.4 ha
Qualche informazione
Isomaltulose (trade names Palatinose and NRGylose)
a disaccharide that is commercially produced
enzymatically from sucrose.
It is a natural constituent of honey and sugar cane and has a very natural sweet
taste. It is particularly suitable as a non-cariogenic sucrose replacement and is
favorable in products for diabetics and prediabetic dispositions.
Like sucrose, it is fully digested and provides the same caloric value of
approximately 4 kcal/g. Unlike sucrose, isomaltulose is toothfriendly and digested
much slower leading not only to a low glycemic response but as well to a
prolonged glucose supply. Thus, isomaltulose is a slowly released carbohydrate
that therefore provides a more sustained energy supply from food and drinks. An
optimal energy supply is a topic of increasing importance in research and product
development as this may play a role in health (particularly control of obesity) as
well as for physical and mental performance.
http://en.wikipedia.org/wiki/Isomaltulose
α-D-glucopyranosyl-(1↔2)-β-D-fructofuranoside
6-0-α-D-Glucopyranosyl-D-fructose
Sucrose
ΔG°′  -12kJ/mol
Isomaltulose
* Not metabolized by many microbes (an acariogenic sweetener)
* Digested by humans with the same primary products (G & F)
* Digested more slowly (lower fluctuation in blood glucose)
Enzyme from Pantoea dispersa UQ68J (91% IM and
3% TH yield from sucrose at 30–35 C)
Enzyme from Pseudomonas mesoacidophila strain
MX-45 (91% TH and 8% IM yield from sucrose at 20°C
Trehalulose
Palatinose
Transgenici
Tuber-specific expression of an apoplasmtargeted SI in potato.
A chimeric sucrose isomerase gene from Erwinia
rhapontici under control of a tuber-specific promoter
was introduced into potato plants. The enzyme
catalyses the conversion of sucrose into palatinose.
Expression of the palI gene within the apoplast of
transgenic tubers led to a nearly quantitative
conversion of sucrose into palatinose.
Despite the soluble carbohydrates having been
altered within the tubers, growth of palI expressing
transgenic potato plants was indistinguishable from
wild type plants.
In growing plant tissues, efficient conversion of sucrose
into the non-metabolized isomer is lethal or disruptive
Fig. 2. HPLC analysis of the soluble
carbohydrates of (A) sugar standards; (B)
extract of potato tubers transformed with palI;
(C) wt potato.
Riduzione del
saccarosio/glucosio
Fig. 3. Contents of palatinose, sucrose, glucose and starch
of wild type potato tubers and transgenic tubers expressing
the chimeric palI gene. Values of wild type (dashed bar)
and transgenic lines (black bars) represent the mean of four
independent measurements ±S.D. As an additional control
transgenic but not expressing lines (n=11) were averaged
and the values are given as mean ±S.D. (grey bar).
glucose (G, 75.0 μM), fructose (F, 37.5 μM), sucrose (S, 37.5
μM), trehalulose (T, 37.5 μM) and isomaltulose (I, 75.0 μM)
High-performance liquid chromatography (HPLC) profiles
from the separation of sugar standards (dotted line) and
4000-fold diluted juice from the basal internode (full lines)
of Q117 control (A) and transgenic line N3.2H (B).
In canna da zucchero
(C) Isomaltulose concentrations in juice
from the basal internodes of transgenic lines.
Transgenic lines with the vacuole-targeted NTPP-68J SI expressed from a
constitutive ubiquitin (Ubi) promoter, were morphologically similar and
equivalent in measured growth parameters to non-transformed controls of the
same background genotype. In contrast, expression of a cytosol targeted form
of the same SI from the same promoter was very damaging to sugarcane plant
development, with low sugar contents and severe stunting in surviving plants.
Controllo
Transgenico N3.2
Transgenico N3.2H
Sugar accumulation profiles of sugarcane cultivar Q117 (A) and transgenic lines N3.2 (B) and N3.2H
(C) showing substantial isomaltulose accumulation and enhanced total sugar concentration in juice.
TVD, top visual dewlap.
Increased photosynthate storage as sugar, rather
than altered partitioning between sugar and fibre.
Total sugar, water and fibre contents per unit fresh weight in internodes of sugarcane
cultivar Q117 control (squares, full line) and SugarBooster transgenic lines N3.2
(triangles, dotted line) and N3.2H (circles, broken line).
Leaf senescence was typically delayed by 15–20 days, resulting in an additional one
to three leaves functional in photosynthesis per stalk for most of the growth period.
Enhanced photosynthesis and sucrose transport in the second fully emerged
leaf (TVD2; TVD, top visible dewlap) of SugarBooster transgenic lines N3.2
and N3.2H, relative to sugarcane cultivar Q117 controls. (A) Photosynthetic
electron transport rate. (B) CO2 assimilation rate.
Trasporto di Saccarosio
(C) Sucrose transport rate into plasma membrane vesicles.
Results are means with standard errors from at least three
replicates. CCCP, carbonyl cyanide m-chlorophenylhydrazone.
Room for improvement?
The vacuole stores a correspondingly large proportion of sucrose, which can
accumulate to 500 μmol/g FW.
Low pH and high protease activity make this a hostile environment to introduced
proteins.
Even in mature stems with substantial IM accumulation, SI enzyme activity was
below the detection limit in cell extracts, consistent with a short half-life of this
protein after delivery into the sucrose storage vacuoles.
Sucrose concentration ceiling?
Mechanisms contributing to the apparent sugar concentration ceiling may
include feedback regulation from broad (e.g. osmotic) or specific (e.g. sucrose)
sensors, thermodynamic limitations, such as leakage of sucrose through storage
compartment membranes, or energetic limitations from the continuous ‘futile’
cycle of sucrose cleavage and synthesis within the storage pool.
Rapid degradation of vacuole targeted SI most likely protects against rapid
sucrose depletion in growing tissues. IM accumulates gradually during
development, probably because of the following: (i) continuous delivery of SI
expressed from the constitutive Ubi promoter; (ii) high catalytic efficiency
allowing occasional IM production before SI inactivation and (iii) absence of
plant enzymes for IM metabolism. For efficient commercial production of this
valued sugar, it will be useful to achieve patterns of developmental expression,
compartmentation and enzyme stability yielding high IM content further up
the harvested stalk profile.
Ingegneria metabolica
probabilmente efficace
(1)
(2)
(3)
(4)
(5)
S
S
S
S
S
A
A
A
A
A
B
B
B
B E
probab. inefficace
In the storage
parenchyma
(6)
(7) (8) cells
(9)of mature
sugarcane stems, the sugar storage vacuole
S of the
S symplast
S
occupiesS about 90%
and
80%+of the total tissue space. The vacuole
stores
TF +a correspondingly large proportion of
A
Ato 500
sucrose,Awhich canAaccumulate
μmol/g
+ FW
B
+
B
B
B
B
Y
Z
W
C
C Q
C F
C
P
P
P Q
P
P
ATP
Q
C
C
C
C
P
P
P
P
X
Lascia o raddoppia?
Doubling the total sugar content in mature internodes of an
elite high-sugar cultivar eliminates osmotic limits and osmotic
sensing as primary constraints behind the previous
concentration ceiling in sugarcane. Our results also challenge
the long-standing assumption that high-sugar genotypes
require at least 70% moisture content in mature internodes.
In the longer term, because sugars ultimately underpin all
other biosynthesis in plants, the SugarBooster effect may be a
foundation for higher yields of many other biomaterials.
Hamerli D, Birch RG (2011) Transgenic expression of trehalulose synthase
results in high concentrations of the sucrose isomer trehalulose in mature
stems of field-grown sugarcane. Plant Biotechnology Journal 9:32-37.
TS transgenic lines yielded up to 350 mM TH without evident reduction in
endogenous sugars. Growth and development of the TH-producing lines were not
visibly different from UbiKN controls at any stage during callus culture, plant
regeneration and growth of the transgenic plants through VG1.
No TH was detectable in juice from mature
internodes of sugarcane background genotype
Q117 or in 35 tested UbiKN control lines
harvested after 12 months of growth in the
glasshouse. TH was detected at this time, at
concentrations up to 542 mM, in 76% of 54
independent transgenic lines that had initially
been selected
Sugars in juice from mature internodes of UBiTS lines in their second (line 2129) or third vegetative
generations in a glasshouse. Each bar represents a single stalk. UniKN control stalks harvested at the
same time yielded 420–705 mM sucrose in juice from mature internodes.
Sugar concentration patterns along the sugarcane stem at maturity in their first field propagation
of (a) UbiKN control line 2141 and (b) UbiTS line 2130 showing substantial isomer production.
Trehalulose concentration in juice increased with internode maturity, reaching about
600 mM, with near-complete conversion of sucrose in the most mature internodes.
Sugars in whole-cane juice extracted at
maturity from UbiTS lines and UbiKN
controls in their first field propagation.
Field performance of transgenic sugarcane
expressing isomaltulose synthase
Basnayake S. (2012) Plant Biotechnology Journal 10:217-225
Developmental profiles of sugar accumulation in stalks
of low-, moderate- and high-level isomaltulose (IM)
producing sugarcane lines.
Relationship between sucrose, total sugars
and isomaltulose (IM) in whole-cane juice
at 10 months in FP2
Developmental profiles of sugar accumulation in stalks
of field-grown recipient genotype Q117 and transgenic
lines N3.2 and N3.2H
Developmental profiles of sugar accumulation in
stalks of recipient genotype Q117 and transgenic line
N3.2 in the field (VG5 at 11 months in May 2007)
and on return to a glasshouse (VG6 at 12 months).
 There was generally a comparable decrease in sucrose concentration,
with no overall decrease in total sugars
 Culture-derived plants were slower to establish and generally gave
shorter and thinner stalks at harvest than those grown from fieldsourced setts in the initial field generations.
 However, after several cycles of field propagation, selections were
obtained with cane yields similar to the recipient genotypes.
 Sucrose isomerase activity was low in these transgenic lines, and the
results indicate strong potential to develop sugarcane for commercialscale production of IM if higher activity can be engineered in
appropriate developmental patterns.
Perchè le stesse linee cresciute in campo non
mostrano più il raddoppio dello zucchero?
I dati nelle diapositive 10 e 21 sono in netto contrasto e non
possono essere spiegati in modo semplice.
Le stesse linee ricresciute in serra e in campo non mostrano
più il fenotipo iniziale e quindi la differenza potrebbe essere
dovuta a dei cambiamenti nelle linee.
 Silenziamento genico?
 Selection for less efficient expressors?
 …?
Chong BF, Bonnett GD, Glassop D, O'Shea MG, Brumbley SM (2007)
Growth and metabolism in sugarcane are altered by the creation of
a new hexose-phosphate sink. Plant Biotechnol J. 5:240-53
Synthesis of sorbitol in sugarcane
using the Malus domestica sorbitol6-phosphate dehydrogenase gene
(mds6pdh).
Rimuovere un prodotto centrale del
metabolismo al momento sbagliato
rischia di compromettere la crescita
(importante la scelta del promotore)
The average amounts of sorbitol detected in the most productive line were 120
mg/g dry weight (equivalent to 61% of the soluble sugars) in the leaf lamina and
10 mg/g dry weight in the stalk pith.
Cosa dire su Sucrose e Sorbitol???
Sugar concentration (mg/g FW) in mature internodes
Sucrose
124.84 ± 12.97
Glucose
1.07 ± 0.36
Fructose
1.21 ± 0.36
(Iskandar (2011) BMC Plant Biology 11:12)
Sorbitol-producing sugarcane generated 30%−40% less aerial biomass and was
10%−30% shorter than control lines.
Leaves developed necrosis in a pattern characteristic of early senescence, and the
severity was related to the relative quantity of sorbitol accumulated.
Section of the last fully expanded leaf from an 8month-old S-76 plant (bottom leaf) and a control
plant (top leaf). Necrosis developed at the S-76 leaf
apex and margins
When the Zymomonas mobilis glucokinase (zmglk)
gene was co-expressed with mds6pdh … the plants
were again smaller, indicating that glucose-6phosphate deficiency was not responsible for the
reduced growth.
Le modificazioni della canna da zucchero sembrano
promettenti sia in termini di aumento di contenuto globale che
in termini di accumulo di composti con maggior valore
rispetto al saccarosio.
Per una effettiva efficacia dal punto di vista commerciale sarà
cruciale modulando l’attività del promotore in maniera da far
produrre l’enzima solo negli internodi più vecchi quando
hanno finito di crescere (quando hanno raggiunto l’effettiva
maturità e non necessitano più saccarosio per crescere).
Aggiungi lavoro su derivato benzoico
Bibliografia
* http://www.ogtr.gov.au/pdf/ir/biologysugarcane08.pdf
* Wu L, Birch RG. (2007) Doubled sugar content in sugarcane plants modified
to produce a sucrose isomer. Plant Biotechnol J. 5:109-17.
* Jackson PJ (2005). Breeding for improved sugar content in sugarcane. Field
Crops Research 92:277–290.
* http://ipmworld.umn.edu/chapters/meagher.htm
* http://www.deutsches-museum.de/en/exhibitions/werkstoffeproduktion/agriculture/sugar-refining/ (Sugarbeet history)
Immagini zuccheri: http://www.chemblink.com/structures/58166-27-1.gif
http://www.3dchem.com/imagesofmolecules/Sucrose.gif
Chong BF, Abeydeera WP, Glassop D, Bonnett GD, O'Shea MG, Brumbley SM
(2010) Co-ordinated synthesis of gentiobiitol and sorbitol, evidence of sorbitol
glycosylation in transgenic sugarcane. Phytochemistry. 71:736-41.
See also ppt "sugarcane in the EAA"
Biancardi E., Campbell L., Skaracis G., De Biaggi M. (eds.) (2005) Genetics
and Breeding of Sugar Beet. Science Publishers Inc., Enfield NH, USA, p.368.
Draycott, A. P (2006) Sugar beet. John Wiley & Sons.