Amaresh Chandra

Expression analysis of genes associated with sucrose accumulation in sugarcane (Saccharum spp. hybrids) varieties differing in content and time of peak sucrose storage.

Sucrose synthesis/accumulation in sugarcane is a complex process involving many genes and regulatory sequences that control biochemical events in source-sink tissues. Among these, sucrose synthase (SuSy), sucrose phosphate synthase (SPS), soluble acid (SAI) and cell wall (CWI) invertases are important. Expression of these enzymes was compared in an early (CoJ64) and late (BO91) maturing sugarcane variety using end-point and qRT-PCR. Quantitative RT-PCR at four crop stages revealed high CWI expression in upper internodes of CoJ64, which declined significantly in both top and bottom internodes with maturity. In BO91, CWI expression was high in top and bottom internodes and declined significantly only in top internodes as the crop matured. Overall, CWI expression was higher in CoJ64 than in BO91. During crop growth, there was no significant change in SPS expression in bottom internodes in CoJ64, whereas in BO91 it decreased significantly. Apart from a significant decrease in expression of SuSy in mature bottom internodes of BO91, there was no significant change. Similar SAI expression was observed with both end-point and RT-PCR, except for significantly increased expression in top internodes of CoJ64 with maturity. SAI, being a major sucrose hydrolysing enzyme, was also monitored with end-point PCR expression in internode tissues of CoJ64 and BO91, with higher expression of SAI in BO91 at early crop stages. Enzyme inhibitors, e.g. manganese chloride (Mn(++) ), significantly suppressed expression of SAI in both early- and late-maturing varieties. Present findings enhance understanding of critical sucrose metabolic gene expression in sugarcane varieties differing in content and time of peak sucrose storage. Thus, through employing these genes, improvement of sugarcane sucrose content is possible.

Sugarcane bud chips: A promising seed material

Sugarcane is normally propagated by stalk cuttings consisting of 2 to 3 bud sett. In conventional system, about 6–8 tons seed cane /ha is used as planting material. Establishing the sugarcane crop using bud chips in place of setts could save about 80% by weight of the stalk material, however this technology has not been scaled up at commercial levels due to poor survival of bud chips under field conditions. Present study is aimed at improving sprouting and establishment of bud chip seed stocks of sugarcane by pre-planting soaking in growth promoting chemicals. Treated bud chips recorded higher bud sprouting, root number, fresh weight of shoot and roots and plant vigor index. Studies have shown that bud chip could be one of the most viable and economical planting material in reducing the cost of sugarcane production.

Exploiting EST databases for the development and characterisation of 3425 gene-tagged CISP markers in biofuel crop sugarcane and their transferability in cereals and orphan tropical grasses

BackgroundSugarcane is an important cash crop, providing 70% of the global raw sugar as well as raw material for biofuel production. Genetic analysis is hindered in sugarcane because of its large and complex polyploid genome and lack of sufficiently informative gene-tagged markers. Modern genomics has produced large amount of ESTs, which can be exploited to develop molecular markers based on comparative analysis with EST datasets of related crops and whole rice genome sequence, and accentuate their cross-technical functionality in orphan crops like tropical grasses.FindingsUtilising 246,180 Saccharum officinarum EST sequences vis-à-vis its comparative analysis with ESTs of sorghum and barley and the whole rice genome sequence, we have developed 3425 novel gene-tagged markers — namely, conserved-intron scanning primers (CISP) — using the web program GeMprospector. Rice orthologue annotation results indicated homology of 1096 sequences with expressed proteins, 491 with hypothetical proteins. The remaining 1838 were miscellaneous in nature. A total of 367 primer-pairs were tested in diverse panel of samples. The data indicate amplification of 41% polymorphic bands leading to 0.52 PIC and 3.50 MI with a set of sugarcane varieties and Saccharum species. In addition, a moderate technical functionality of a set of such markers with orphan tropical grasses (22%) and fodder cum cereal oat (33%) is observed.ConclusionsDeveloped gene-tagged CISP markers exhibited considerable technical functionality with varieties of sugarcane and unexplored species of tropical grasses. These markers would thus be particularly useful in identifying the economical traits in sugarcane and developing conservation strategies for orphan tropical grasses.

Physio-Biochemical and Molecular Approaches to Enhance Sucrose Content in Sugarcane: Indian Initiatives

Sucrose is the prime product of sugarcane hence continued research is needed to improve its level in cane stalk. Bio-physiological aspects with relation to growth and sucrose storage mechanism are still poorly understood though the morphology and anatomy of sugarcane has been extensively studied. Still too many gaps exist in our knowledge as far as unique development, physiology and synthesis/degradation of sucrose in sugarcane are concerned. It is imperative to first visualize the impact of a selected trait on general sugarcane physiology as crop is highly compensatory in nature. Eventually, these gaps have slow down the precise improvement to enhance the expression of desired agronomical traits. Increased photosynthetic capacity of variety concomitant with increased cane yield has no value unless until sugar content is increased in that proportion. Realistically, it is difficult to increase sucrose level in cane stalk without prior knowledge of the factors that affect sugarcane yield and carbon partitioning. Since many genes involved in the metabolism of sucrose including signals and factors responsible for its regulation, further complicates the situation. Factors related to multiple environmental constraints namely temperature, water deficit and excess, increased CO2 and availability of nutrient directly or indirectly influence or cross talk the controlling pathways of growth and sucrose metabolism. In the present review an attempt has been made to discuss and explore the possibility of enhancing sucrose productivity in sugarcane addressing the issues pertaining to physiology, biochemistry and biotechnology primarily focusing India as the active centre for the sugar and sugarcane production.

Expression analysis of genes associated with sucrose accumulation in sugarcane under normal and GA3-induced source–sink perturbed conditions

Sugarcane accumulates high amount of sucrose, thus making it one of the important cash crops worldwide. The final destination of sucrose accumulation in sugarcane is sink tissue, i.e., stalk, supplied by the source, i.e., leaf, to fulfill the need of plant growth, respiration, storage, and other metabolic activities. Signals between sink and source tissues regulate sucrose accumulation in sink and possibly the negative feedback from the sink restrains further accumulation in the stalk. However, perturbation of this negative feedback may help to improve sugar yield. This can be achieved by the application of GA3 (Gibberellic acid), a plant growth regulator, known to excite physiological responses and modify the source–sink metabolism through their effect on photosynthesis, which in turn improves sink strength by redistribution of the photoassimilates. In the present study, GA3 applied canes showed prominent increase in invertase activity, at early stage of the application, to provide hexoses. This in turn helped increase the internodal length and cane capacity for additional accumulation of sucrose, thereby increasing sink strength. At maturity, sucrose% and brix% were found higher in middle and top portions of the GA3-applied canes. Expression analysis of various sucrose metabolising genes viz., sucrose phosphate synthase (SPS), sucrose synthase (SuSy), soluble acid invertase, neutral invertase, and cell wall invertase (CWI) was carried out at different growth stages, using quantitative RT-PCR. CWI, which plays key role in phloem unloading in sink tissues, exhibited higher expression in GA3 samples at the elongation stage which decreased with maturity, whereas both SuSy and SPS, involved in regulation of sucrose accumulation, showed a variable level of expression. Thus, GA3 application on cane may improve the sucrose content in stalk and thus assuage maneuvering source–sink dynamics in sugarcane.

Biochemical Profiling of Source and Sink Tissues at Different Growth Stages of Early and Late Maturing Varieties of Sugarcane (Saccharum spp. hybrids)

Sugarcane being C4 crop exhibits distinct source-sink signaling pathway that helps in storing remarkably high amount of sucrose in its sink tissues that makes it a highly remunerable crop worldwide. In the present study sugar content was profiled in both source and sink tissues of early (CoJ64) and late (BO91) maturing sugarcane varieties. At early growth stage (i.e. at 210 DAP) sink tissues of both varieties exhibited higher reducing sugar and low sucrose content while in source tissues both sucrose and reducing sugar content was observed high, depicted lower sink demand for sucrose. With maturity, when sink demand for sucrose storage increased, rise in sucrose content was seen in sink tissues, whereas in source tissues gradual decrease in sucrose and reducing sugar content was observed. Accumulation of sucrose was found much higher in CoJ64 than those in BO91. In CoJ64 maximum sucrose content (64.2%) was seen at 330 DAP while in BO91 it was 41.8% at 390 DAP. At this stage, source tissues too exhibited higher sucrose and reducing sugar content. Thus sucrose synthesis in source tissues and its transportation to the sink tissues is primarily governed by the sink demand.

Biotechnology of Stylosanthes

The genus Stylosanthes contains some 40 species largely classified on morphological characteristics and has a monophyletic origin, closely related to Arachis with the S. guianensis species complex as the most ancient group clearly distinct from the rest of the genus. Many species are important forage legumes and grown both in annual and perennial system of agriculture. Species are grouped in two subgeneric sections, sections Styposanthes and Stylosanthes based on the presence or absence, respectively, of an axis rudiment, a small appendage at the base of the pod or loment. Most species of Stylosanthes are diploid (2n = 20) but polyploid species (2n = 40 and 2n = 60) also exist. Latter are exclusively allopolyploid. Section Styposanthes contains both diploid and polyploid species, while species in Section Stylosanthes are exclusively diploid. Molecular markers have proved that a tertraploid (4n) is a combination of a diploid (2n) species from section Stylosanthes and a diploid (2n) species from Sec. Styposanthes. A reasonable numbers of sequence-tagged site (STS) markers have been developed in this genus and used to define the progenitors of tetraploid S. scabra, S. hamata and only hexaploid species S. erecta. Genetic diversity study in this genus is largely based on isozyme, RAPD and STS markers. Lack of sufficient EST sequences and simple sequence repeat (SSR) markers have hampered a desired good linkage map; however, RAPD and few RFLPs based map delivered QTLs for drought and anthracnose disease. Before the introduction of Stylosanthes seabrana in year 1998–1999, S. scabra, S. hamata, S. guianensis and S. humilis were major species cultivated in Indian tropics and sub-tropics. Though regeneration protocols have been developed for few species, gene transfers through genetic engineering means have been very limited in this genus. Incorporation of leaf meal, stylo meal (as the replacements of concentrate) in feeding system of small and large ruminants, poultry ration as well as their conservation in form of fodder banks are some of the new and impacted initiatives have been taken with Stylosanthes crop in South Asian countries. Narrow genetic base, insufficient SSR markers, loose genetic map and overall low breeding efforts world wide in Stylosanthes invite concerted attention for its improvement by exploiting genomic resources and information being generated with related legumes, vital to this less explored but an important forage legume for better and improved livestock husbandry.

Effect of leaf stripping on cane and sugar yield in sugarcane

Effect of leaf stripping +2 levels at monthly intervals starting from August till December on cane yield and commercial cane sugar (CCS) of sugarcane was studied under field conditions. Treatments comprised of whole leaf removal (WLR) and leaf blade removal (LBR). Cane yield showed significant improvement when stripping was performed in the month of August and November and the magnitude was greater with leaf blade removal; 26.1 % and 17.9 % over control in the month of August (LBR) and November (WLR), respectively. CCS % was numerically higher in most of the stripping treatments. Stripping significantly increased CCS (t/ha) in some of the treatments (August, LBR, November WLR and December LBR). The Leaf Area Index (LAI) showed maximum reduction (55.7% and 62%) in the month of November. Results obtained indicated that the effect of LBR was more beneficial than WLR for improving cane and sugar yield and also stripped leaves could be used as animal fodder after 7 month of crop age without affecting sugarcane yield.