Sugarcane Breeding Programs Research Articles

A quantitative genetics approach to nitrogen use efficiency in sugarcane

The economic and environmental consequences of inefficient use of nitrogen (N) fertiliser in agricultural crops is of concern worldwide, so new crop varieties with improved nitrogen use efficiency (NUE) are sought. Here, we report the first study of mapping quantitative trait loci (QTL) for nitrogen physiology traits in sugarcane. QTL analysis was undertaken for each parent of a segregating bi-parental sugarcane mapping population. We grew 168 progeny under limiting (0.2 mM NH4NO3) and non-limiting (5.0 mM NH4NO3) N supplies in two glasshouse experiments. Significant marker-trait associations (MTA) were detected in each treatment for shoot dry weight, root dry weight, total shoot N, shoot internal NUE (iNUE; measured as units shoot dry weight per unit N), leaf protein content and glutamine synthetase (GS) activity. MTA for GS activity did not co-locate with other traits except leaf protein content, indicating that variation in GS activity is not linked to plant size or iNUE during early growth. Under high N, there were no significant MTA for iNUE among markers from the male parent, Q165, an Australian commercial cultivar, but six MTA were found for markers inherited from the female parent, IJ76–514, a Saccharum officinarum ancestral variety. The results indicate that variation for iNUE under high N may be lower in commercial varieties than unimproved genotypes. Further, four MTA were consistent with previous field-based research on sugar and biomass production. Our study provides initial evidence that QTL may be incorporated in sugarcane breeding programs targeting improved NUE.

SSR marker-based analysis of genetic relatedness among sugarcane cultivars (Saccharum spp. hybrids) from breeding programs in China and other countries

Capillary electrophoresis-based molecular genotyping was conducted on 35 sugarcane cultivars (Saccharum spp. hybrids) and 5 clones of related wild species with 20 polymorphic SSR DNA markers. A total of 251 alleles were identified with 248 alleles displaying varying degrees of polymorphism and the remaining three alleles being monomorphic. The total number of alleles by any SSR marker varied from as few as 7 to as many as 18, with an average of 12.5 alleles per marker. Diversity index (DI = 1 − \( \sum\limits_{i = 1}^S {Pi^2 } \)) for these SSR markers ranged from 0.71 to 0.91, with a mean of 0.83. A composite parameter NDI, representing a product of the number of alleles (N) and DI, is an indicator on the general usefulness of a DNA marker. Ten SSR markers, namely, mSSCIR43, mSSCIR66, SMC119CG, SMC24DUQ, SMC278CS, SMC31CUQ, SMC336BS, SMC597CS, SMC703BS, and SMC851MS, have NDI values of greater than 12 in comparison to less than 10 from the rest markers, indicating that these 10 SSR markers provide much information for genotyping the 40 clones. A finding to minimize stutters and minus-Adenine peaks gave a guideline for the selection of best SSR markers for other SSR research. The 35 cultivars were clustered into five groups based on pairwise similarity coefficient values and their relationships to the wild species were demonstrated. Inclusion of CP67-412, CP72-1210, N21, N27, and S. officinarum clone Badila into the cultivar groups is due to the fact that these clones have been extensively used as parental material in sugarcane breeding programs. The results are in general agreement with the evolutionary course of the sugarcane cultivars that the order of contributing species in modern sugarcane cultivars is S. officinarum, S. spontaneum, S. robustum Brandes et. Jesw.ex., S. sinense Roxb., and S. barberi Jesweit. The accordance of molecular results with recorded evolution of sugarcane verified the fidelity and usefulness of these 20 SSR markers in progeny selection and allele transmission study in this aneu-polyploidy crop.

Impact of depth of pedigree and inclusion of historical data on the estimation of additive variance and breeding values in a sugarcane breeding program

Sugarcane breeders in Australia combine data across four selection programs to obtain estimates of breeding value for parents. When these data are combined with full pedigree information back to founding parents, computing limitations mean it is not possible to obtain information on all parents. Family data from one sugarcane selection program were analysed using two different genetic models to investigate how different depths of pedigree and amount of data affect the reliability of estimating breeding value of sugarcane parents. These were the parental and animal models. Additive variance components and breeding values estimated from different amounts of information were compared for both models. The accuracy of estimating additive variance components and breeding values improved as more pedigree information and historical data were included in analyses. However, adding years of data had a much larger effect on the estimation of variance components of the population, and breeding values of the parents. To accurately estimate breeding values of all sugarcane parents, a minimum of three generations of pedigree and 5 years of historical data were required, while more information (four generations of pedigree and 7 years of historical data) was required when identifying top parents to be selected for future cross pollination.

Repeatability Between Two Intermediate Sugarcane Genotype Selection Stages in Florida

Improved yield and disease resistance on sand soils are priorities of the Canal Point (CP) sugarcane (Saccharum spp.) breeding and selection program. Analyses of historical phenotypic data can provide helpful information in guiding selection strategies to meet these priorities. Correlation analysis was used to examine repeatability of phenotypic data used to advance genotypes from an unreplicated single location clonal crop test (stage II) to the subsequent stage (stage III; two replicate, four location clonal crop experiment). Correlations between data for four traits measured in stage II and the corresponding data pooled across soil types for the same genotypes in stage III varied across 23 series of the CP program. Generally, when correlations were statistically significant (P < 0.05), correlation values were low (means; theoretical recoverable sucrose (TRS) r = 0.40, cane yield r = 0.27, and economic index r = 0.23). Similar trends were evident for correlations between data from stage II and stage III on muck soil and stage II and stage III on sand soil across 10 series of the CP program. A 10% reduction in the number of genotypes advanced to stage III over that period would have meant losing only 1 and 13 genotypes that had commercial potential on muck and sand soils, respectively (n = 1278). Correlations between the phenotypic data were significant only for stage III comparisons between TRS and cane yield, which were negatively associated on either soil type. These results indicate that changes in the advancement strategy from stage II are not required as advancing approximately 135 genotypes identifies almost all genotypes with the genetic potential to yield well on muck or sand soils in stage III. Increasing genotypes in stages prior to stage III and changing crossing strategies to improve identification of disease-resistant, high-yielding genotypes for sand soils is recommended.

The Brazilian experience of sugarcane ethanol industry

Biomass has gained prominence in the last few years as one of the most important renewable energy sources. In Brazil, a sugarcane ethanol program called ProAlcohol was designed to supply the liquid gasoline substitution and has been running for the last 30 yr. The federal government’s establishment of ProAlcohol in 1975 created the grounds for the development of a sugarcane industry that ­currently is one of the most efficient systems for the conversion of photosynthate into different forms of energy. Improvement of industrial processes along with strong sugarcane breeding programs brought technologies that currently support a ­cropland of 7 million hectares of sugarcane with an average yield of 75 tons/ha. From the beginning of ProAlcohol to the present time, ethanol yield has grown from 2,500 to around 7,000 l/ha. New technologies for energy production from crushed sugarcane stalk are currently supplying 15% of the electricity needs of the country. Projections show that sugarcane could supply over 30% of Brazil’s energy needs by 2020. In this review, we briefly describe some historic facts of the ethanol industry, the role of sugarcane breeding, and the prospects of sugarcane biotechnology.

Proline Accumulation, Photosynthetic Abilities and Growth Characters of Sugarcane ( Saccharum officinarum L.) Plantlets in Response to Iso-Osmotic Salt and Water-Deficit Stress

The aim of this study was to investigate the biochemical, physiological and morphological responses of sugarcane to iso- osmotic salt and water-deficit stress. Disease-free sugarcane plantlets derived from meristem cuttings were photo- autotrophically grown in MS media and subsequently exposed to −0.23 (control), −0.67 or −1.20 MPa iso-osmotic NaCl (salt stress) or mannitol (water-deficit stress). Chlorophyll a (Chl a), chlorophyll b (Chl b), total carotenoids (C x+c), maximum quantum yield of PSII (Fv/Fm), photon yield of PSII (ΦPSII), stomatal conductance (Gs) and transpiration rate (E) in the stressed plantlets were significantly reduced when compared to those of plantlets of the control group (without mannitol or NaCl), leading to net-photosynthetic rate (Pn) and growth reduction with positive correlation. In addition, physiological changes and growth parameters of plantlets in the salt stress conditions were more sharply reduced than those in water- deficit stress conditions. On the other hand, the proline content and non-photochemical quenching (NPQ) in the leaves of stressed plantlets increased significantly, especially in response to iso-osmotic salt stress. The chlorophyll pigments in iso-osmotic stressed leaves were significantly degraded ( r 2 = 0.93), related to low water oxidation ( r 2 = 0.87), low netphotosynthetic rate ( r 2 = 0.81), and growth reduction ( r 2 = 0.97). The multivariate biochemical, physiological and growth parameters in the present study should be further used to develop salt, or drought, tolerance indices in sugarcane breeding programs.

Relative performance of the sugarcane families in early selection stages

Family selection is widely used in early stages of selection in sugarcane breeding programs. Families are evaluated in the first one or two stages of selection and individual clones are selected from within superior families. These selected clones are evaluated for their performance in two or three selection stages before commercial release of superior clones. This study was undertaken to assess the relative performance of five sugarcane families for predicting potential parental combinations and also to identify rare and elite recombinants that can be exploited in future. Results from this study highlighted that two families viz., Co 7704 x Co 8209 and Co 85002 x Co 86011 contributed a greater proportion of elite clones combining acceptable levels of cane yield and sugar yield. The top ten clones selected based on sugar yield from the stage II evaluation holds greater promise in the subsequent regional selection programs. Overall, the findings suggest that selection of the best families based on their mean performance and further selection of individual clones based on their sugar yield in early stages would improve the efficiency of selection and increase heritability in the genetic populations being tested.

Location Contributions Determined via GGE Biplot Analysis of Multienvironment Sugarcane Genotype‐Performance Trials

Selection for productive sugarcane (Saccharum spp.) cultivars in Florida has been more successful for organic than for sand soils. The objectives of this study were to assess the contributions of a sand‐soil location to the final stage of multienvironment testing of sugarcane genotypes in Florida, and to identify locations with organic soils that, if replaced with a sand‐soil location, would be least likely to compromise superior cultivar selection for organic soils in Florida. Sixteen genotypes were harvested in two or three crop cycles from 2002 to 2005 at nine locations. Traits analyzed were cane and sucrose yields (Mg ha−1) and theoretical recoverable sucrose (TRS) (g kg−1). The sand‐soil location, Lykes, was generally neither highly representative of locations nor highly discriminating of genotypes. Results revealed the desirability of replacing an organic‐soil location with a sand‐soil location in the final testing stage of this sugarcane breeding and selection program. Caution must be exercised, however, to ensure that such action would not compromise genotype discrimination for TRS and sucrose yield. Ability to identify productive cultivars on organic soils by the Florida sugarcane selection program would be least compromised by replacing either Osceola or Knight with a sand‐soil location.

Management of ash/impurity ratio in sugarcane: relative effects of genotypes, and N and K fertiliser rates

In sugarcane juice or raw sugar, ash refers to soluble inorganic salts. The ratio of ash/impurity is important because high levels reduce sucrose extraction in sugar mills and the market value of raw sugar. The aim of research reported here was to examine, through a series of field experiments, the effects of genotypes and varying N and K fertiliser application rates on ash and ash/impurity levels. Ash levels were estimated via conductivity measurements, following prior studies indicating close relationships between conductivity and ash. Significant variation due to genotypes for conductivity/impurity levels was observed, with a range of about ±25% around the mean. Despite this variation, broad-sense heritability of conductivity/impurity levels on unselected genotypes on the basis of measurements in one environment were low (0.20), suggesting that selection in early phases of selection systems in breeding programs would probably be of limited value. However, it was recommended that measurements on conductivity/impurity ratio be made in multi-environment trials for genotypes being tested for commercial release, to assist in comprehensive economic evaluation. On average it was found that conductivity/impurity levels increased by 15% per 100 kg/ha of applied K, highlighting potential costs of high rates of K fertiliser. Increased N rates had a significant but small effect on reducing conductivity/impurity levels by ~10% per additional 100 kg/ha of applied N. The information reported here may be used to help develop optimal strategies for cultivar selection in sugarcane breeding programs and for developing recommendations for optimal fertiliser management, and some guidelines in relation to this are discussed.

Polyphasic characterization of Gluconacetobacter diazotrophicus isolates obtained from different sugarcane varieties.

A polyphasic approach was applied to characterize 35 G. diazotrophicus isolates obtained from sugarcane varieties cultivated in Brazil. The isolates were analyzed by phenotypic (use of different carbon sources) and genotypic tests (ARDRA and RISA–RFLP techniques). Variability among the isolates was observed in relation to the carbon source use preference. Glucose and sucrose were used by all isolates in contrast to myo-inositol, galactose and ribose that were not metabolized. The results of the analysis showed the presence of two groups clustered at 68% of similarity. The genetic distance was higher when RISA-RFLP analysis was used. Analysis of 16S rDNA sequences from isolates showed that all of them belonged to the G. diazotrophicus species. Neither effect of the plant part nor sugarcane variety was observed during the cluster analysis. The observed metabolic and genetic variability will be helpful during the strain selection studies for sugarcane inoculation in association with sugarcane breeding programs.

Genetic diversity associated with in vitro and conventional bud propagation of Saccharum varieties using RAPD analysis

AbstractPolymorphisms in the genomic DNA of eight varieties maintained by conventional bud propagation (via rhizomes) and by in vitro shoot tip cultures were detected by RAPD analysis of sugarcane varieties. The study estimated the genetic diversity induced after in vitro multiplication of these varieties. Higher (28.9%) and lower (12%) numbers of polymorphic bands were detected in plants propagated via rhizomes; the genetic similarity estimate varying from 0.63 to 0.80. Plants of SP90‐3723 and SP91‐1049, or RB85‐5113 and SP90‐3723, varieties involving greater genetic distances may be indicated as progenitors in breeding programmes. In vitro multiplication of RB86‐7515, RB85‐5113, RB83‐5054 and SP86‐42 varieties increases genetic variability, while in vitro multiplication of SP91‐1049, SP90‐1638, RB92‐8064 and SP90‐3723 leads to genetic similarity. Results show that the RAPD technique is an effective tool for detecting polymorphism in sugarcane clones and it allows quick collection of the necessary information (more genetically divergent plant varieties) to guide new crossing in sugarcane breeding programmes.

Sucrose‐Phosphate Synthase, a Biochemical Marker of High Sucrose Accumulation in Sugarcane

Increasing sucrose content is a major objective of sugarcane breeding programs. One approach employed by breeders is to introgress new genes from genotypes of Saccharum officinarum L. not previously used in breeding. The activity of a suite of key sucrose metabolizing enzymes was measured in progeny of an introgression program to find biochemical markers associated with high sucrose content, measured as commercial cane sugar (CCS). The enzymes sucrose‐phosphate synthase (SPS), three isoforms of invertase, and sucrose synthase were measured in four high and four low CCS clones from an initial cross between a S. officinarum and the commercial cultivar Q165. Subsequently, SPS and the two soluble isoforms of invertase were measured in clones derived from a backcross of one of the progeny to another commercial cultivar Mida. Enzyme activities were measured in tissue from internodes taken from four different positions down the stem profile. Of particular significance was the finding that the activity of a key enzyme involved in sucrose synthesis, SPS, was significantly higher in the upper internodes (one to three) of high CCS clones as compared with low CCS clones in both populations, suggesting that this enzyme may have a key role in establishing metabolic and developmental processes necessary for high sugar accumulation during stem growth and maturation.

Varietal selection for perennial crops where data relate to multiple harvests from a series of field trials

Varietal selection for yield from a series of multi-environment trials can be regarded as a multi-trait selection problem in which the yields in different environments are synonymous with traits. As such an analysis of the data combined across environments should be conducted in order to form an index for selection. Analytical methods that include appropriate models for both the genetic variance structure (that is, the variances and covariances of genotype effects from different environments) and the residual variance structure (which typically comprises spatial covariance models for each trial) have been published previously. In the case of perennial crops, yields are often obtained from multiple harvests which implies that the data comprise short sequences of repeated measurements. Varietal performance in individual harvests is important for selection so that a combined analysis across both trials and harvests is required. The repeated measures nature of the data provides additional modelling challenges. In this paper we propose an approach for the analysis of multi-environment, multi-harvest data that accommodates the major sources of variation and correlation (including temporal). The approach is illustrated using two examples from sugarcane breeding programmes. The proposed models were found to provide a superior fit to the data and thence more accurate selection decisions than the common practice of conducting separate analyses of individual trials and harvests.

Characterization of intergeneric hybrids of Erianthus rockii and Saccharum using molecular markers

Erianthus rockii, a wild relative of sugarcane, is drought and cold tolerant, and both are potentially important agronomic traits to the sugarcane industry worldwide. As such it is of interest as a source of parental germplasm to sugarcane breeders and is currently being used in sugarcane introgression programs in both China and Australia. To date morphological characters have been used to verify the putative hybrids produced. Two crosses were generated between two different Saccharum species and two E. rockii accessions. Over 400 AFLP markers were used to identify the intergeneric hybrid progeny as well as determine hybrid diversity. Both crosses generated hybrids but efficiency levels were very different and are probably related to the different Saccharum parent used in each cross. Cross 1 was between a Saccharum officinarum and E. rockii and generated 100% hybrid progeny. Cross 2, however, was between a sugarcane hybrid (S. officinarum × Saccharum spontaneum) and E. rockii and only 10% of the progeny were intergeneric hybrids. Inheritance of markers in the progeny was analysed and for both crosses there were equal numbers of markers from both parents indicating n + n transmission of chromosomes. This is the first verification of E. rockii hybrids with molecular markers. It may now be possible to exploit genes of value from E. rockii in sugarcane breeding programs.

Associations between DNA markers and resistance to diseases in sugarcane and effects of population substructure

Association between markers and sugarcane diseases were investigated in a collection of 154 sugarcane clones, consisting of important ancestors or parents, and cultivars. 1,068 polymorphic AFLP and 141 SRR markers were scored across all clones. Data on the four most important diseases in the Australian sugarcane industry were obtained; these diseases being pachymetra root rot (Pachymetra chaunorhiza B.J. Croft & M.W. Dick), leaf scald (Xanthomonas albilineans Dowson), Fiji leaf gall (Fiji disease virus), and smut (Ustilago scitaminea H. & P. Sydow). By a simple regression analysis, association between markers and diseases could be readily detected. However, many of these associations were due to the effects of embedded population structure and random effects. After taking population structure into account, we found that 59% of the phenotypic variation in smut resistance ratings could be accounted for by 11 markers, 32% of variation for leaf scald and pachymetra root rot rating by 4 markers, and 26% of Fiji leaf gall by 5 markers. The results suggest that marker-trait associations can be readily detected in populations generated from modern sugarcane breeding programs. This may be due to special features of past sugarcane breeding programs leading to persistent linkage disequilibrium in modern parental populations.

Conservation ofSaccharum spontaneum as defuzzed true seed

Storing true seed (fluff) of sugarcane germplasm at low temperatures for preserving its germinability was highlighted as an alternative or complementary method for their maintenance and further utilisation in a Sugarcane Breeding Programme. The viability of defuzzed true seed collected from 50Saccharum spontaneum clones stored at -20°C temperature for five years was investigated .The germination potential of the seed was evaluated annually using germination tests. It was found that the seed of the germplasm could be stored safely without much reduction in germinability. The practical advantage of maintaining sugarcane germplasm as defuzzed true seed under cold storage conditions has been highlighted.

Effects of plot size on the estimation of genetic parameters in sugarcane families

An optimization of the use of resources in sugarcane breeding programs that would not interfere with the genetic evaluations is desirable and of great importance. The costs could be reduced and the chances of success would be higher through the possibility of investing rationally in a larger number of available genetic resources. Our study evaluated the smallest number of rows per plots of sugarcane families for yield traits; the variations in the parameter estimates for different plot sizes were discussed; and the ideal plot size was estimated by the maximum curvature method. Plot reduction leads to gradual CV increases and interferes with the ratio CVg/CVe, which in some cases dropped to values below one unit, indicating experimental inaccuracy. Heritability estimates decrease with plot size. The maximum curvature method showed that 2-row plots are enough to evaluate all characteristics.

Effect of Genotype and Genotype × Nitrogen Rate Interactions on Color in Juice and Raw Sugar from Sugarcane

There is an increased emphasis being placed on raw sugar quality in international sugar markets. Color intensity of raw sugar is one quality parameter of importance, with high color levels increasing refining costs and hence lowering sugar value. The quality of raw sugar is compromised by colored solutes or precursors of colored compounds. We examined genetic effects on color levels in juice from sugarcane (Saccharum spp.) in three field experiments, two of which also included different N fertilizer rate treatments. The relative impact on amino N in juice, an important precursor of colored compounds, was also examined. A key finding was consistently large variation among varieties (approximately threefold differences) for color and color‐to‐impurity ratio levels. These effects were highly repeatable across different environments and N rates, with genotype × environment and genotype × N rate interaction variance being not significant or less than 15% of genotype main effects in all experiments. Relatively small genetic effects were detected for amino N levels and these were less repeatable across environments than for color. Thus, selection for low color levels will be effective in sugarcane breeding programs. A relative color index is proposed to compare genotypes for propensity to produce color in raw sugar, considering the effects of both color and amino N in juice.

AFLP Analysis of the Phenetic Organization and Genetic Diversity in the Sugarcane Complex, Saccharum and Erianthus

Amplified fragment length polymorphism (AFLP) markers were evaluated for determining the phylogenetic relationships, and the diversity in the Saccharum complex using 30 clones belonging to S. officinarum, S. robustum, S. spontaneum, S. barberi, S. sinense and the related genus Erianthus. The phenetic tree of the species clones based on AFLP data was consistent with the known taxonomical relationships. AFLP gave higher resolution of closely related species into discrete groups than that by RAPD and RFLP markers, reported earlier. The levels of diversity within the various Saccharum species were also found to be higher than those obtained previously with the same set of clones using RAPD markers. The intraspecies similarity in S. barberi and S. sinense was much higher than interspecies similarity suggesting a clear separation of the two, which are considered ‘horticultural species’. The genetic similarity matrix derived from a single primer combination highly correlated (r = 0.980) with that obtained from all the 12 primer combination used in the study, thus highlighting the efficiency of a single primer combination in delineating species relationships. All the primer combinations could identify markers that are specific to each of the species and the genus Erianthus. Among the species, specific markers were highest in S. spontaneum followed by S. robustum, S. barberi, S. officinarum and S. sinense. Erianthus had a distinct profile with 30% of the total amplified fragments being specific to it. This offers great scope for identifying intergeneric hybrids, which has been very difficult using morphological traits and RAPD markers. High degree of correspondence between the results from the cluster analysis based on Jaccard's similarity index, Neighbour Joining tree based on Sokal and Michener distance matrix and AFTD (Analyses Factorielle on Table of Distances) analysis clearly demonstrated that AFLP markers would be an appropriate tool in providing better information about the relationships among the species, estimation of diversity, and in revealing species and genus specific markers that could be directly applied in sugarcane breeding programmes.

Analysis of Genetic Diversity Among Saccharum spontaneum L. From Four Geographical Regions of India, Using Molecular Markers

Saccharum spontaneum L. a wild relative of sugarcane is the most variable and diverse among the Saccharum species. This species had been successfully exploited in sugarcane improvement programmes since 1915 and most of the present day commercial varieties are derivatives of interspecific hybrids involving S. spontaneum. The S. spontaneum germplasm available today in the World collections is diverse and represent different geographical groups. In the present investigation, an attempt was made to characterize 40 S. spontaneum clones collected from 4 different geographical areas in India using 20 random, 2 ISSR and 2 telomere primers. Of the 491 bands generated by these primers 83.9% were polymorphic. The genetic diversity estimated based on these markers was found to be moderate (48.9%). The pair-wise genetic distance between the accessions ranged from 29.8 to 60.0. The accessions from Arunachal Pradesh were found to be the most diverse, while Tamil Nadu accessions showed relatively less diversity. Diversity between Tamil Nadu and Kerala collections was found to be low, while the diversity between the Orissa group and the rest was found to be high. The collections from Mayurbanj and Cuttack regions of Orissa were found to be distinct. Arunachal Pradesh accessions, being more diverse, are a potential source for exploitation in sugarcane breeding programmes.