C. L. McIntyre

Identification of genomic regions associated with stay green in sorghum by testing RILs in multiple environments

Abstract Stay green is an important drought resistance trait for sorghum production. QTLs for this trait with consistent effects across a set of environments would increase the efficiency of selection because of its relatively low heritability. One hundred and sixty recombinant inbreds, derived from a cross between QL39 and QL41, were used as a segregating population for genome mapping and stay green evaluation. Phenotypic data were collected in replicated field trials from five sites and in three growing seasons, and analysed by fitting appropriate models to account for spatial variability and to describe the genotype by environment interaction. Interval mapping and non-parametric mapping identified three regions, each in a separate linkage group, associated with stay green in more than one trial, and two regions in single trial. The regions on linkage groups B and I were both consistently identified from three trials. The multiple environment testing was very helpful for correctly identifying QTLs associated with the trait. The utilisation of molecular markers for stay green in sorghum breeding is also discussed.

A combination of AFLP and SSR markers provides extensive map coverage and identification of homo(eo)logous linkage groups in a sugarcane cultivar

Sugarcane varieties are complex polyploids carrying in excess of 100 chromosomes and are derived from interspecific hybridisation between the domesticated Saccharum officinarum and the wild relative S. spontaneum. A map was constructed in , an Australian cultivar, from a segregating F1 population, using 40 amplified fragment length polymorphism (AFLP) primer combinations, five randomly amplified DNA fingerprints (RAF) primers and 72 simple sequence repeat (SSR) primers. Using these PCR-based marker systems, we generated 1,365 polymorphic markers, of which 967 (71%) were single-dose (SD) markers. Of these SD 967 markers, 910 were distributed on 116 linkage groups (LGs) with a total map length of 9,058.3 cM. Genome organisation was significantly greater than observed in previously reported maps for Saccharum spp. With the addition of 123 double-dose markers, 36 (3:1) segregating markers and a further five SD markers, 1,074 markers were mapped onto 136 LGs. Repulsion phase linkage detected preferential pairing for 40 LGs, which formed 11 LG pairs and three multi-chromosome pairing groups. Using SSRs, double-dose markers and repulsion phase linkage, we succeeded in forming 127 of the 136 LGs into eight homo(eo)logy groups (HG). Two HGs were each represented by two sets of LGs. These sets of LGs potentially correspond to S. officinarum chromosomes, with each set aligning to either end of one or two larger LGs. The larger chromosomes in the two HGs potentially correspond to S. spontaneum chromosomes. This suggestion is consistent with the different basic chromosome number of the two species that are hybridised to form sugarcane cultivars, S. spontaneum (x=8) and S. officinarum (x=10), and illustrates the structural relationship between the genomes of these two species. The discrepancy of coverage between HGs highlights the difficulty in mapping large parts of the genome.

Prediction of hybrid performance in grain sorghum using RFLP markers

Abstract.Heterosis is an important component of hybrid yield performance. Identifying high yielding hybrids is expensive and involves testing large numbers of hybrid combinations in multi-environment trials. Molecular marker diversity has been proposed as a more efficient method of selecting superior combinations. The aim of this study was to investigate the value of molecular marker-based distance information to identify high yielding grain sorghum hybrids in Australia. Data from 48 trials were used to produce hybrid performance-estimates for four traits (yield, height, maturity and stay green) for 162 hybrid combinations derived from 70 inbred parent lines. Each line was screened with 113 mapped RFLP markers. The Rogers distances between the parents of each hybrid were calculated from the marker information on a genome basis and individually for each of the ten linkage groups of sorghum. Some of the inbred parents were related so the hybrids were classified into 75 groups with each group containing individual hybrids that showed similar patterns of Rogers distances across linkage groups. Correlations between hybrid-group performance and hybrid-group Rogers distances were calculated. A significant correlation was observed between whole genome-based Rogers distance and yield (r = 0.42). This association is too weak to be of value for identifying superior hybrid combinations. One reason for the generally poor association between parental genetic diversity and yield may be that important QTLs influencing heterosis are located in particular chromosome regions and not distributed evenly over the genome. Variation in the sign and magnitude of correlations between Rogers distance and hybrid-group performance for particular linkage groups observed in this study support this hypothesis. The concept of using diversity on individual linkage groups to predict performance was explored. Using data from just two linkage groups 38% of the variation in hybrid performance for grain yield could be explained. A model combining phenotypic trait data and parental diversity on particular linkage groups explained 71% of the variation in grain yield and has potential for use in the selection of heterotic hybrids.

Assessing genetic diversity in a sugarcane germplasm collection using an automated AFLP analysis

An assessment of genetic diversity within and between Saccharum, Old World Erianthus sect. Ripidium, and North American E.giganteus (S.giganteum) was conducted using Amplified Fragment Length Polymorphism (AFLPTM) markers. An automated gel scoring system (GelComparTM) was successfully used to analyse the complex AFLP patterns obtained in sugarcane and its relatives. Similarity coefficient calculations and clustering revealed a genetic structure for Saccharum and Erianthus sect. Ripidium that was identical to the one previously obtained using other molecular marker types, showing the appropriateness of AFLP markers and the associated automated analysis in assessing genetic diversity in sugarcane. A genetic structure that correlated with cytotype (2n=30, 60, 90) was revealed within the North American species, E. giganteus (S.giganteum). Complex relationships among Saccharum, Erianthus sect. Ripidium, and North American E.giganteus were revealed and are discussed in the light of a similar study which involved RAPD markers.

Quantitative trait loci identified for sugar related traits in a sugarcane (Saccharum spp.) cultivar × Saccharum officinarum population

AbstractThe identification of markers linked to quantitative trait loci (QTLs) for increased sugar accumulation could improve the effectiveness of current breeding strategies in sugarcane. Progeny from a cross between a high sucrose producing cultivar, (denotes Australian plant breeding rights), and a Saccharum officinarum clone, IJ76-514 were grown in two field experiments in different years, and evaluated in the early and mid-season phases of crop maturity, to identify robust QTLs in affecting sucrose content in cane. Using an extensive genetic map constructed for with over 1,000 AFLP and SSR markers, a total of 37 QTLs were identified for brix and pol of which, 16 were detected in both experiments. Of these 37 QTL, 30 were clustered into 12 genomic regions in six of the eight homo(eo)logous groups. Each QTL explained from 3 to 9% of the phenotypic variation observed. Both positive and negative effects were identified and the location of the QTLs on linkage groups belonging to the same homo(eo)logy group suggested that a number of the QTLs were allelic forms of the same genes. Of the 37 QTLs identified, the majority were significant in both early and mature cane, but 8 were identified as early specific QTLs and 9 as mature cane QTLs. In total, 97 interactions were significant (P<10−5) and these were localised to 32 genomic regions of which 6 were detected with both years’ data. Models including all the QTLs explained from 37 to 66% of the total phenotypic variation, depending on the trait. The results will be subsequently applied in marker assisted breeding.

Identifications of two different mechanisms for sorghum midge resistance through QTL mapping.

Sorghum midge is the one of the most damaging insect pests of grain sorghum production worldwide. At least three different mechanisms are involved in midge resistance. The genetic bases of these mechanisms, however, are poorly understood. In this study, for the first time, quantitative trait loci associated with two of the mechanisms of midge resistance, antixenosis and antibiosis, were identified in an RI (recombinant inbred) population from the cross of sorghum lines ICSV745 × 90562. Two genetic regions located on separate linkage groups were found to be associated with antixenosis and explained 12% and 15%, respectively, of the total variation in egg numbers/spikelet laid in a cage experiment. One region was significantly associated with antibiosis and explained 34.5% of the variation of the difference of egg and pupal counts in the RI population. The identification of genes for different mechanisms of midge resistance will be particularly useful for exploring new sources of midge resistance and for gene pyramiding of different mechanisms for increased security in sorghum breeding through marker-assisted selection.

Analysis of phylogenetic relationships in the triticeae tribe using RFLPs

The use of restriction fragment length polymorphisms in combination with other approaches is very useful for the reconstruction of evolutionary events revealing phylogenetic relationships. A set of 21 cDNA probes hybridizing to different chromosome arms in hexaploid wheat was used in a series of experiments designed to estimate the phylogenetic relationships among and within 16 species of the Triticeae tribe. A high degree of polymorphism was found both between and within the species examined. The RFLP data were used to generate a cladogram and a phenogram in order to compare the two different methods of constructing phylogenetic trees. The results of both methods were consistent with each other and with the general taxonomic information provided by earlier morphological studies, meiotic pairing analysis, isozyme tests, and sequence alignment in theTer,NOR, and5s DNA loci. In addition, several correlations were found between the geographical origin of accessions from the same species and their phylogenetic relationships as shown by the cladogram and phenogram.

Construction of a genetic linkage map for Saccharum officinarum incorporating both simplex and duplex markers to increase genome coverage

Saccharum officinarum L. is an octoploid with 80 chromosomes and a basic chromosome number of x = 10. It has high stem sucrose and contributes 80% of the chromosomes to the interspecific sugarcane cultivars that are grown commercially for sucrose. A genetic linkage map was developed for S. officinarum (clone IJ76-514) using a segregating population generated from a cross between Q165 (a commercial sugarcane cultivar) and IJ76-514. In total, 40 AFLP and 72 SSR primer pairs were screened across the population, revealing 595 polymorphic bands inherited from IJ76-514. These 595 markers displayed a frequency distribution different from all other sugarcane genetic maps produced, with only 40% being simplex markers (segregated 1:1). Of these 240 simplex markers, 178 were distributed on 47 linkage groups (LGs) and 62 remained unlinked. With the addition of 234 duplex markers and 80 biparental simplex markers (segregating 3:1), 534 markers formed 123 LGs. Using the multi-allelic SSR markers, repulsion phase linkage, and alignment with the Q165 linkage map, 105 of the 123 LGs could be grouped into 10 homology groups (HGs). These 10 HGs were further assigned to the 8 HGs observed in cultivated sugarcane and S. spontaneum. Analysis of repulsion phase linkage indicated that IJ76-514 is neither a complete autopolyploid nor an allopolyploid. Detection of 28 repulsion linkages that occurred between 6 pairs of LGs located in 4 HGs suggested the occurrence of limited preferential chromosome pairing in this species.

Ribosomal DNA variations in Erianthus, a wild sugarcane relative (Andropogoneae-Saccharinae)

Variation at the 18S+26S and 5S ribosomal DNA loci was assessed on 62 Erianthus Michx. clones, representing 11 species, and 15 clones from two Saccharum L. species used as a reference. Genus-specific markers for Erianthus Michx. sect. Ripidium Henrard (Old World species) were identified. Ribosomal DNA units in Erianthus sect. Ripidium exhibited an additional BamHI site compared to Saccharum, and 5S units showed length and restriction-site differences between Erianthus and Saccharum. These markers will be useful to follow introgression in Saccharum x Erianthus hybrids. Six ribosomal units (for 18+26S genes) were revealed in Erianthus sect. Ripidium, differing by restriction-site positions and/or length. These results provided new information on species relationships and evolution within the genus Erianthus. The Indonesian and Indian forms of E. arundinaceus (Retz.) Jeswiet gave different restriction patterns, which were similar to those of E. bengalense (Retz.) R. C. Bharadwaja and E. procerus (Roxb.) Raizade, respectively. The two 2n=20 species, E. ele-phantinus Hook.f. and E. ravennae (L.) P. Beauv., could also be differentiated at this locus. Two of the New World Erianthus species studied, E. rufipilus (Steud.) Griseb. and E. longisetosus Andersson, appeared more like Erianthus sect. Ripidium, whereas E. trinii Hack, and E. brevibardis Michx. showed patterns consistent with Miscanthus sinensis Andersson and S. spontaneum L., respectively. Finally, the comparison of rDNA restriction maps among Erianthus sect. Ripidium, Saccharum, sorghum and maize, led to unexpected conclusions concerning the relationships between the different genera and the position of Erianthus in the “Saccharum complex”.

Loss of genetic diversity associated with selection for resistance to sorghum midge in Australian sorghum

In recent years, hybrids with levels of resistance to sorghum midge (Stenodiplosis sorghicola Coquillett) have become available to Australian sorghum producers. These hybrids have been readily accepted to the extent that more than 80% of the sorghum growing area was planted to hybrids with some level of midge resistance by 1995. Since selection for resistance to sorghum midge is one of the primary objectives of Australian sorghum breeding programs, the relationship between resistance and genetic diversity was investigated.Genetic diversity and heterozygosity were assessed using restriction fragment length polymorphism analysis among 26 grain sorghum hybrids grown commercially in Australia.The genetic distances between each sorghum hybrid and a standard highly resistant hybrid were found to be strongly negatively correlated to hybrid midge resistance ratings (r = - 0.77, p < 0.001). In addition, the average heterozygosity of each hybrid was correlated with their midge resistance ratings (r = - 0.54, p < 0.01).The results indicate that the move to midge resistant hybrids has been associated with a narrowing of the genetic diversity and average heterozygosity of commercial sorghum hybrids. Repeated use of particular elite parent lines, linkage drag and genetic drift are likely to have contributed to this decline. This reduction in genetic diversity may have implications for the genetic vulnerability of sorghum in Australia and the rate of progress in breeding for yield.