Thesis (MSc (Genetics))--University of Stellenbosch, 2007. / The aim of this study was to develop a DNA-based diagnostic system that can be used to distinguish between genotypes in the wheat breeding program at the University of Stellenbosch. Known marker systems were investigated and the chosen marker system would then be implemented to determine its utility in the breeding program. Three marker systems were considered, i.e. microsatellites, Amplified Fragment Length Polymorphisms (AFLPs) and various retrotransposon-based markers. Each system is based on polymerase chain reaction (PCR) amplification from specific primer pairs. The multitude of primer options was narrowed down during a review of published literature regarding wheat molecular markers. Thirty nine microsatellite primer pairs and nine AFLP primer combinations were chosen for the initial genotype evaluation. Four different retrotransposonbased techniques were investigated; namely Inter-Retrotransposon Amplified Polymorphism (IRAP), REtrotransposon-Microsatellite Amplified Polymorphism (REMAP), Sequence- Specific Amplified Polymorphism (SSAP) and, a derivative of these developed in this study, Wis-2 Retrotransposon Amplification. The study started with twenty genotypes which included varieties/breeding lines from five breeding programmes. The genotypes were chosen as representative of the respective breeding populations and were used in the initial testing of the marker systems. Eighteen microsatellites were evaluated using the panel of twenty genotypes. From this, six primer pairs (Xgwm190, Xgwm437, Xgwm539, Xwmc11, Xwmc59 and Xwmc177) were chosen to test the semi-automated DNA sequencer detection system. A single band/peak in each microsatellite profile was used for genotyping. Four of the primer pairs were labelled with different fluorochromes which enabled them to be multiplexed. The differences in amplification products of the six microsatellites meant that all six could be detected in one electrophoresis run. The banding pattern produced by microsatellite Xwmc177 was complex and highly polymorphic and was therefore also analysed in the same way as the AFLP patterns. When analyzed in this manner it proved to be more informative than the combination of six microsatellites (with a single prominent band scored in each). Three AFLP primer combinations could also be multiplexed and visualised together. The three EcoRI selective primers were labelled with different dyes and used with one MseI selective primer. The SSAP system also used fluorescently labelled primers and proved to be the most useful of the retrotransposon-based methods. However, this system produced such a large amount of data that it made analysis too time consuming. Therefore the six microsatellites and three AFLP primer combinations (MseI-CTC and EcoRI-ACA, -AAC, - AGG) were selected for routine genotyping. Due to the numerous highly polymorphic bands produced by the SSAP system it could be very useful to differentiate very closely related genotypes that cannot be distinguished with the markers proposed for routine use. A panel of 119 breeding lines were then used to implement the two chosen marker systems. The results obtained for these markers were used to produce a dendrogram of the lines using the SAS cluster analysis function. The clusters showed that most of the lines could be distinguished from each other. The MseI-CTC and EcoRI-AGG primer combination was the most informative. It produced the largest number of clusters (53) and could therefore discriminate between more of the lines than any other method. The dendrograms and clusters allowed sixteen of the breeding lines to be selected to test the optimal number of seeds to represent an entire population (variety/breeding line) as one seed was not sufficient. It was decided that eight seeds could provide a good representation of the intra-line variability.

Thesis (MScAgric.)--University of Stellenbosch, 2001. / ENGLISH ABSTRACT: In cross pollinated crops, recurrent selection is used to increase the frequency of desirable alleles by breaking up existing linkage blocks and forming new gene combinations. Despite promising results from numerous feasibility studies, recurrent selection is seldom routinely used in wheat. A major obstacle has been the inability to readily achieve random interbreeding of large numbers of selected plants. In China the Taigu genetic male sterility gene, Ms2, has however been used to establish a recurrent selection programme in which field grown male sterile plants were pollinated by selected male fertile plants (Huang et al., 1988). Another dominant gene for male sterility, Ms3, was found after EMS treatment of the seeds of an alloplasmie common wheat with Triticum tauschii cytoplasm (Maan et al., 1984) and is located at 3 map units from the centromere on chromosome arm SAS (Maan et al., 1987). In a study done during 1999 at Welgevallen to determine the frequency of natural intererossing under field conditions, Ms3 showed incomplete penetrance and only about two thirds of the seed set on male sterile plants could be attributed to intercrossing. Ms3 has stable expression in plants grown within the normal range of greenhouse temperatures for wheat, 16 - 2SoC. Under warmer field conditions, 21 - 3SoC, its penetrance is, however, incomplete (Maan et al., 1984). The utility of Ms3 under field conditions is therefore unsatisfactory. An attempt to determine the location and origin of an unknown male sterility gene, found in cross 9SK3 of a routine breeding programme, showed that a single locus was not the cause of the male sterility. Chromosome abnormalities and gene imbalances were probably to blame. The male sterility probably relates to a T.urartu addition chromosome in the pedigree of cross 9SK3. To facilitate the production of large numbers of hybrid progeny, a simple hydroponic system was developed in which male sterile tillers cut at the flowering stage can be pollinated and maintained for about 8 weeks, long enough to produce viable seeds. For pollination, florets on male tillers are cut open and placed in a container with a similar number of pollen shedding male tillers. It was found that cut tillers could be maintained in the hydroponic system as long as certain precautions were met: (a) The tillers must be handled with care so as not to damage the flag leaf which must be maintained for as long as period possible. (b) The tillers have a nutrient requirement and a 20% solution showed the best results of the nutrient solutions tested. (c) The sterilizing effect of Jik at O.OS%gave excellent fungal control en helped to sustain the nutrient solution. (d) Although the treatment of tillers with hormones improved seed quality, it was not justified by the additional inputs required. Different selection strategies were used for male and female plants. At the onset of the recurrent selection programme in 1998, a total of 1881 plants were tested for seedling resistance and 597 plants were selected for use as parents and source material for 1999. In total 158 male sterile and 188 male fertile ears were used in the hydroponic pollination and a 63.47% seed set was obtained, resulting in 3410 seeds, forming the 1999 female component. One hundred and fifty seven F2:96K109plants were selected from a field grown population in 1998. These, together with 44 selections from a pedigree programme, formed the male component for 1999. In total 9564 plants were tested for seedling resistance during 1999. A total of 3230 resistant seedling were selected and planted. Again male fertile plants from the previous season were field planted and selected. The selected plants were subjected to mixograph testing. A total of 448 male sterile and 1020 male fertile ears were used for hydroponic pollination. Approximately 12000 seeds were harvested, the seed set being around 75%. The 157 F2:96K109 field selected plants (1999) and 64 selections from a pedigree programme formed the male component for 2000. Seedling resistance testing during 2000 included a total of 6465 plants and 2832 were selected and planted. The hydroponic system was improved during 2000 with new, larger capacity containers being used which improved cross pollination. In total 878 male sterile tillers and 1016 male fertile tillers were cut and intercrossed. In total 25380 seeds were harvested, the seed set being 81.7%. In an attempt to determine the amount of variation within the 157 F2-families selected during 1999, mixograph testing was performed. The data showed variation among families. Seedling resistance testing for leaf and stem rust was performed on the 1999 and 2000 FIs to determine the variation for resistance within the populations. Both populations showed high level of stem rust resistance but lower levels of leaf rust resistance (± 50%). Ms3 can thus be used in combination with hydroponic tiller culture to facilitate recurrent selection. Integration with an excisting pedigree selection programme is viable and requires little additional input. Some of the these results have already been published (Addendum D). / AFRIKAANSE OPSOMMING: Herhalende seleksie word by kruisbestuiwers aangewend om die frekwensie voordelige allele te verhoog deur die opbreek van bestaande koppelingsblokke en vorming van nuwe geen-kombinasies. Hoewel uitstekende resultate m.b.V.herhalende seleksie reeds by koring verkry is, is die roetine aanwending egter beperk weens die gebrek aan effektiewe kruisbestuiwing van groot getalle plante. In China is "Taigu" genetiese manlike steriliteit, Ms2, egter met sukses aangewend vir die vestiging van 'n herhalende seleksieprogram vir landverboude koring. Die manlik-vrugbare plante word vir die bestuiwing van geselekteerde manlik-steriele plante aangewend (Huang et al., 1988). Nog 'n dominante manlike steriliteitsgeen, Ms3, is ontdek na EMS behandeling van sade afkomstig vanaf 'n alloplasmiese gewone koring met 'n Triticum tauschii sitoplasma (Maan et al., 1984) en is gesetelop chromosoom 5AS, 3 kaarteenhede vanaf die sentromeer (Maan et al., 1987). 'n Ondersoek na die frekwensie natuurlike kruisbestuiwing onder landtoestande (Welgevallen, 1999) het getoon dat onvolledige penetrasie van Ms3 lei tot ongeveer 5% selfbestuiwing en dat slegs twee-derdes van die saadset aan kruisbestuiwing toegeskryf kon word. Ms3 word wel stabiel uitgedruk onder normale glashuistemperature tydens blom nl. 16 - 25°C, maar onder warmer landtoestande, 21 - 35°C, is uitdrukking onstabiel met laer penetrasie van die geen (Maan et al., 1984). Die benutbaarheid van Ms3 onder landtoestande was dus onbevredigend. Die ondersoek na die oorsprong en ligging van 'n onbekende, manlike steriliteitsgeen (95K3) wat ontdek is in 'n roetine teelprogram het daarop gedui dat 'n enkellokus waarskynlik me ter sprake is nie, maar eerder chromosoom-abnormaliteite en geenwanbalanse. Die manlike steriliteit kan verband hou met 'n T urartu addisie chromosoom in die stamboom van hierdie bron. Ten einde kruisbestuiwing van 'n groot aantal plante te bewerkstellig, is 'n eenvoudige bestuiwersisteem ontwikkel gegrond op waterkultuurkweking van afgeknipte manlik-steriele (Ms3ms3), are. Manlik-steriele en manlik-vrugbare are is tydens blom geknip. Die manliksteriele are se blommetjies is oopgeknip en toegelaat om deur die manlik-vrugbare are bestuif te word. Die bestuifde manlik-steriele are (Ms3ms3) is hierna vir ongeveer 8 weke gelaat vir saadvorming. Afgeknipte are kan baie suksesvol in voedingsmedium onderhou word mits sekere eenvoudige voorsorgmaatreëls getref word, naamlik: (a) Die are moet met sorg hanteer word en die vlagblaar moet so lank as moontlik behou word. Are moet weekliks teruggeknip word ten einde verstopping en agteruitgang van vaatweefsel teen te werk. Die oorspronklik- afgeknipte halm is dus belangrik. (b) Die are toon 'n definitiewe voedingsbehoefte en 'n 20% voedingsoplossing was die beste van die oplossings wat getoets is. Die voedingsoplossing moet verkieslik weekliks vervang word wanneer are teruggeknip word. Op die tydstip behoort die houers met 'n steriliseringsmiddel gewas te word vir die verwydering van enige moontlike swamgroei aan die houers se wande. (c) Jik was die beter steriliseringsmiddel en het teen 0.05% toediening goeie swaminhibering bewerkstellig. (d) Hormone is nie in die roetinetoepassing gebruik nie aangesien die voordeel hiervan nie die ekstra insette regverdig nie. Verskillende strategieë is aangewend vir die seleksie van manlike en vroulike plante. Met die aanvang van die herhalende seleksieprogram in 1998 is 'n totaal van 1881 plante getoets vir roesweerstand en 597 geselekteer as bronmateriaal vir 1999. In totaal is 158 manliksteriele en 188 manlik-vrugbare are gebruik in die bestuiwersisteem vir die verkryging van die 1999 vroulike komponent. 'n Totaal van 3410 sade is verkry met 'n 63.47% saadset. Tesame met 157 F2:96KI09 landgeselekteerde plante is 44 seleksies vanuit 'n stamboom seleksieprogram gebruik as manlike komponent in 1999. Gedurende 1999 is 9564 plante getoets vir roesweerstand en 3230 geselekteer en geplant. Weereens het landseleksie plaasgevind. Die 157 seleksies is onderwerp aan miksograaf-toetsing. Vierhonderd agt- en - veertig manlik-steriele en 1020 manlik-vrugbare are is gebruik in die bestuiwersisteem. Ongeveer 12138 sade is geoes, teen 'n 75% saadset. Gedurende 2000 is die sade asook 64 seleksies uit 'n stamboom seleksieprogram aangewend as die manlike komponent. Roestoetsing is weereens in 2000 uitgevoer en 6465 plante is geïnokuleer waaruit 2832 plante geselekteer en geplant is. Die bestuiwersisteem is aangepas vir die hantering van groter aantalle are tydens 2000 en in totaal is 878 manlik-steriele are en 'n 1016 manlik-vrugbare are gebruik vir kruisbestuiwing. Die saadset is verhoog na 81.7% en 25380 sade is verkry. Om die hoeveelheid variasie binne die populasie te bepaal, is miksograaftoetsing op die 1999 F2-populasie uitgevoer. Die data het aangetoon dat groot hoeveelhede genetiese variasie beskikbaar is binne die populasie. Roestoetsing van die 1999- en 2000-bestuiwerpopulasies is ook uitgevoer om 'n indikasie te verkry van die verspreiding van weerstand teen blaar- en starnroes. Die blaamoes het 'n relatief lae vlak van weerstand getoon (± 50%) terwyl die stamroesweerstand baie hoë vlakke gehandhaaf het. Ms3 kan dus gebruik word om in kombinasie met waterkultuurkweking van gesnyde halms, 'n herhalende seleksieprogram van stapel te stuur. Integrasie met 'n bestaande stamboom seleksieprogram is ook moontlik en sal relatief min addisionele insette vereis. 'n Gedeelte van die werk is reeds gepubliseer en word hierbyaangeheg as Aanhangsel D.

Please read the summary in the section 02back (Appendix 1) of this document / Thesis (PhD (Genetics))--University of Pretoria, 2005. / Genetics / unrestricted

Thesis (PhD)--Stellenbosch University, 2015 / ENGLISH ABSTRACT : Diuraphis noxia Kurdjumov (Russian Wheat Aphid; RWA) is a pest of wheat and barley that has spread from its home range in the fertile crescent to most wheat producing countries except Australia. Since its first introduction to South Africa and the USA in the late 20th century, breeding programs for wheat phenotypes resistant to the aphid were put in place. Conventional breeding practices rely on phenotypic screening to verify traits carried by offspring and genetic tools such as marker assisted selection (MAS) have greatly aided this process in speed and accuracy. The size and complexity of the wheat genome, its allopolyploid nature and repetitive elements have, however, posed a challenge to studies on the genetics of this cereal crop. Many studies have focused on chromosome 3B which is the largest of the wheat chromosomes and easily separated from the redundant genomic background by techniques such as flow cytometry. The similarity in size of the remaining chromosomes however, limits the application of flow cytometry to their isolation. Databases such as Grain-Genes (http://wheat.pw.usda.gov/GG2/index.shtml) house marker data from various mapping studies for all wheat chromosomes and in 2014 the International Wheat Genome Sequencing Consortium (IWGSC) completed the draft genome sequence of wheat categorized by chromosome. Sources of resistance (Dn resistance genes) against RWA are located on chromosome 7D. but despite the marker and sequence data available currently, mapping studies specific for the Dn resistance genes are few. Additionally, sequence data available is derived from cultivars susceptible to RWA and is not comprehensively annotated and assembled in many cases. In this study, we demonstrate a novel, combined approach to isolate and characterize the Dn resistance genes through the use of a genetic map constructed from Amplified Fragment Length Polymorphism (AFLP), Expressed Sequence Tag (EST) and microsatellite markers and a physical map constructed from Next Generation Sequencing (NGS) data of ditelosomic chromosomes (7DS and 7DL) isolated by microdissection on the PALM microbeam system. A 122.8 cM genetic map was produced from 38 polymorphic AFLP markers and two ESTs with the microsatellite Xgwm111 as anchor to related genetic maps. Through comparison to maps available on GrainGenes the location of the Dn1 resistance gene was narrowed down to a deletion bin (7DS5-0.36-0.62) on the short arm of chromosome 7D with an AFLP marker (E-ACT/M-CTG_0270.84) mapping closely at 3.5 cM and two ESTs mapping at 15.3 cM and 15.9 cM from Dn1. Isolation of individual chromosome arms 7DS and 7DL using the PALM Microbeam system allowed sequencing of the chromosome without the redundancy of the remainder of the hexaploid genome. Through isolating the chromosome arms in this way, a >80-fold reduction in genome size was achieved as well as a major reduction in repetitive elements. Analysis of the sequencing data confirmed that 7DL is the physically shorter arm of the chromosome though it contains the majority of protein coding sequences. / AFRIKAANSE OPSOMMING : Diuraphis noxia Kurdjumov (Russiese koring-luis; RWA) is « pes wat op koring en gars voorkom. Die pes het vanaf sy tuiste in die midde Ooste na meeste koringproduserende lande behalwe Australië versprei. Sedert die eerste bekendstelling van RWA in Suid Afrika en die VSA in die vroeë 20ste eeu is teelprogramme ten gunste van koring lyne met weerstand teen RWA begin. Tradisionele teelprogramme maak op fisieise observasie van die fenotipe staat om te verifieer of plante in die nageslag die gewenste eienskap dra. Genetiese metodes soos merkerondersteunde seleksie (MAS) versnel hierdie selekteringsproses grootliks. Die grootte en kompleksiteit van die koring genoom asook die polyploïde en herhalende natuur daarvan is « groot hindernis vir genetiese studies van hierdie graangewas. Baie studies het op chromosoom 3B gefokus wat die grootste van die koring chromosome is en dus maklik vanaf die res van die oorbodige genomiese agtergond deur tegnieke soos vloeisitometrie geskei word. Die ooreenkoms in grootte tussen die res van die chromosome bemoeilik die toepassing van vloeisitometrie om hulle te isoleer. Databasisse soos GrainGenes (http://wheat.pw.usda.gov/GG2/index.shtml) bevat merker data vanaf verskeie karterings-studies vir al die chromosome en in 2014 het die "International Wheat Genome Sequencing Consortium"(IWGSC) die voorlopige basispaarvolgorde van die koring genoom bekendgestel, gekategoriseer volgens chromosoom. Weerstandsbronne (Dn weerstandsgene) teen RWA kom meestal op chromosoom 7D voor. Ten spyte van merker en basispaarvolgorde data tans beskikbaar is karterings-studies spesifiek tot die Dn gene skaars en basispaarvolgorde data is vanaf kultivars afkomstig wat nie weerstandbiedend teen RWA is nie en waarvan die annotasie en samestelling baie keer nie goed is nie. In hierdie studie demonstreer ons « nuwe, gekombineerde aanslag om die Dn weerstandsgene te isoleer en karakteriseer deur van « genetiese kaart opgestel met "Amplified Fragment Length Polymorphism"(AFLP), "Expressed Sequence Tag"(EST) en mikrosatelliet merkers asook « fisiese kaart saamgestel deur die volgende-generasiebasispaarvolgordebepaling van ditelosomiese chromosome (7DS en 7DL) geïsoleer deur mikrodisseksie met die "PALM Microbeam"sisteem gebruik te maak. « Genetiese kaart van 122.8 cM was met 38 polimorfiese AFLP merkers en twee EST merkers geskep. Die mikrosatelliet, Xgwm111, is ook ingesluit en het as anker vir verwante genetiese-kaarte gedien. Deur vergelyking met genetiese-kaarte op GrainGenes is die posisie van die Dn1 weerstandsgeen vernou na « delesie bin (7DS5-0.36-0.62) op die kort arm van chromosoom 7D met « AFLP merker (EACT/ M-CTG_0270.84) wat ongeveer 3.5 cM vanaf die geen karteer. Die twee EST merkers is 15.3 cM en 15.9 cM vanaf die geen gekarteer. Isolering van die individuele chromosoom arms, 7DS en 7DL, deur van die "PALM Microbeam"sisteem gebruik te maak het basispaarvolgordebepaling van die chromosoom toegelaat sonder die oortolligheid van die res van die hexaploïde genoom. Deur die chromosoom so te isoleer is « >80-maal verkleining in genoom grootte bereik insluitend « groot reduksie in herhalende elemente. Analise van die data vanaf basispaarvolgordebepaling het bevestig dat chromosoom 7D die fisiese kleiner chromosoom is maar dat dit die meerderheid van proteïn koderende basispaarvolgordes bevat.

Weed infestation is one of the yield limiting factors in crop production. Weeds have negative effect on crop growth and productivity due to competition, allelopathy or hosting other harmful organisms. For large-scale wheat production, the use of wide spectrum pre-emergence or post-emergence herbicides remains the most valuable weed control tool. In South Africa, annual grass weeds are a major wheat production constraint, which is usually managed through application of pre-emergence herbicides. Due to limited water availability and low soil moisture content, these herbicides can often become ineffective and result into high weed infestations, which then have to be managed by manual cultivation or post-emergence herbicidal applications. However, there are no effective selective post-emergence herbicides available to control grass weeds in wheat. There is also limited option to use broad-spectrum post-emergent herbicides because they non-selectively kill the crop and weeds. Consequently, the use of herbicide resistant crops is a viable weed management system in wheat production. Breeding herbicide resistant crop varieties would allow farmers to safely use post-emergence herbicides without damaging the crop. Subsequently yield and quality losses will be reduced significantly. Thus, the development of herbicide resistant crop varieties through mutation breeding is a novel approach for effective weed management under both small-scale and commercial farmers. Mutagenesis has been recognized as one of the most efficient method to induce genetic variation in plants. Through induced mutations, development of new variants is possible that could be manipulated in plant breeding programs. Mutation leads to alteration of various traits in crop plants including plant height, improved nutritional quality, shorter growing period, increased tolerance or resistance to abiotic and biotic stresses. Ethylmethanesulphonate (EMS) is one of the most widely used chemical mutagens to induce mutagenesis in crop plants. The objectives of this study were to: 1) determine the optimum EMS concentration, treatment temperature and duration that would provide desired germination percentage and vigorous and healthy seedlings for effective mutagenesis in wheat, 2) investigate variations in agro-morphological traits in two selected wheat varieties (SST56 and SST875) after EMS mutagenesis and 3) select herbicide resistant wheat germplasm after inducing genetic variation using EMS using two selected wheat varieties (SST56 and SST875). The objectives were achieved through three independent studies as outlined below: In the first study seeds of four selected wheat varieties (B936, B966, SST387 and SST875) were treated in two replicates with three EMS concentrations (0.3, 0.5, and 0.7%), three temperature regimes (30, 32.5 and 35 °C) at four time durations (0.5, 1, 1.5 and 2 hrs). Results showed highly significant interactions (P<0.01) among varieties, EMS concentrations, temperature and exposure time on seedling emergence, germination and seedling height. Seeds treated with the highest EMS dose (0.7%), temperature (35ºC) and long exposure time (2 hr) showed delayed emergence by 18 days. At 30ºC, 0.5hr and 0.3% EMS varieties B936, B966 and SST875 had early emergence (6 days). B936 and SST387 had 50% while B966 and SST875 had 53% and 57% germination, respectively. These results were observed at EMS level of 0.7%, 300C and 1.5 hr exposure time in B936 and EMS at 0.5%, 350C and 1.5 hr in B966. SST387 and SST875 required EMS dose at 0.5%, 32.50C and 2 hr treatment time. Other low or high treatment combinations were invariably ineffective comparedto untreated control. During the second study two selected varieties (SST56 and SST875) were subjected to EMS mutagenesis using 0.5% v/v EMS at 32.5oC for 1 hr. Field trials were carried out at Ukulinga research farm of the University of KwaZulu-Natal in the randomized complete block design with two replicates. Data on nine important agro-morphological traits were collected and analyzed using the analysis of variance (ANOVA), correlation and principal component analysis (PCA) procedures. Significant variations were found among the agro-morphological traits between M1 individuals compared to untreated checks. The mutagenesis significantly reduced seed germination in the field at 40% in both varieties. The treatment significantly delayed days to heading by 8 days and shortened days to maturity by 13 days in both varieties. EMS treatment also significantly reduced plant height at 18 cm in SST56 and 21 cm in SST875 and spike length reduced by ~2.5 cm in both varieties. Plant height had positive and significant correlation with number of tillers, number of seeds per spike, flag leaf length and 100 seed weight. However, it had negative correlation with the number of days to maturity. The PCA revealed that three principal components (PC1, PC2 and PC3) accounted to 57% of the total variations among the agro-morphological traits in both varieties. PC1 alone contributed to 27.7% of the variation which was well-correlated with plant height (0.767), tiller number (0.812), number of seeds per spike (0.599) and seed yield (0.720). PC2 explained 15.6% of the variation and well-correlated with germination percentage (0.784), spike length (0.554) and flag leaf length (0.772). PC3 accounted to 12.4% of the variation and had negative correlation with days to maturity (-0.730). In the last study, seeds of two selected wheat varieties (SST56 and SST875) were treated with EMS at 0.5% concentration for 2 hr at 32.5ºC. Treated seeds and comparative controls were planted at the experimental farm of the University of KwaZulu-Natal using the randomized complete block design. Four weeks after planting M1 plants and untreated standard checks were sprayed with two herbicides, i.e. metsulfuron-methyl and bromoxynil at three different doses viz. 2x, 4x and 8x above the recommended rate of 4 g ha-1 and 2 kg ha-1, respectively. Two weeks after the treatment herbicide resistance were assessed. Results showed significant difference among varieties, tested herbicides and doses used. The EMS treated wheat lines showed variable degree of herbicide resistance compared to untreated controls. Overall, the study established the requirement of variety specific EMS dose and treatment temperature and duration that could be used for inducing large-scale mutation to select targeted mutant individuals in wheat. Further, the study found that EMS has the potential to increase agro-morphological variations in wheat to select useful and novel mutants with desired phenotypic traits and herbicide resistance which will be subjected for further selections to identify stable and herbicide resistance lines. / Thesis (M.Sc.Agric.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

Gaeumannomyces tritici is a soil-borne, highly destructive, wheat root pathogen, causing take-all disease. Some modern, elite, winter wheat cultivars possess a genetic trait promoting low take-all inoculum build-up (LowTAB). This leads to reduced disease if wheat is grown in the same field the next year. This PhD aimed to test if genetics (LowTAB) and chemistry will individually or synergistically influence take-all fungal inoculum build-up in first wheats as methods to control second wheat take-all disease. The underlying mechanism, epidemiology, agronomy and genetics of the TAB (take-all build-up) trait in eight first wheat field trials was investigated. This identified two minor QTLs conferring the LowTAB trait, in a doubled haploid mapping population. This PhD also confirms the highly complex cultivar-year-field interactions that underpin this trait. Root phenotyping experiments in the field and laboratory highlight that the TAB trait is not likely to be the result of root system architecture variation. Future field trials are planned to confirm the QTLs identified and to test for links between TAB and root-soil-microbial interactions. The effect of foliar applied chemistry (fungicide: Amistar, active ingredient: azoxystrobin and plant growth regulator: Moddus, a.i. trinexapac-ethyl) combined with genetics (TAB) on first wheat take-all inoculum build-up and second wheat disease was investigated. To complement this, laboratory screens were performed checking for common target site mutations to the azoxystrobin fungicide, in new and historic G. tritici isolates. For the first time, legacy effects of first wheat foliar chemistry on second wheat disease were identified, however no synergy with genetics were found. Early first wheat Amistar sprays reduced second wheat take-all disease, whilst later sprays and plant growth regulator, Moddus; had no effect. However, first wheat inoculum reduction by Amistar, could not be directly linked to the second wheat disease outbreaks observed. No evidence of fungicide resistance was found in 40 UK isolates, thus the varied efficacy of Amistar is linked to soil dose rate at the different application times. The collective PhD findings of the effect of first wheat chemistry and genetics make a significant contribution to the control of take-all disease in commercial second wheat crops.

Thesis (MSc (Genetics))--Stellenbosch University, 2008.

[Truncated abstract] Wheat is the main crop in Australia and there are stringent quality requirements. Preharvest sprouting induced by rainfall between maturity and harvest lowers grain quality from premium to feed grades and reduces yield. Wheat production has expanded into the southern Western Australian region where preharvest sprouting occurs in ~1 in 4 seasons and development of more preharvest sprouting tolerant genotypes is required. The main mechanism for improving preharvest sprouting tolerance is grain dormancy. There is genetic variation for dormancy based in the embryo and seed coat but dormancy is complex and is influenced by environmental conditions during grain filling and maturation. Screening and selecting for preharvest sprouting tolerance is problematic and the level of tolerance needed for regions which differ in the level of dormancy they impose, requires clarification. The research presented here aims to answer the underlying question for breeders of how much dormancy is required for preharvest sprouting tolerance in contrasting target environments of the central and coastal wheat belt regions of Western Australia. In the central and coastal wheat belt regions, field trials with modified environments were used to determine the environmental influence on dormancy. Water supply (without directly wetting the grain) and air temperature were modified during grain development in a range of genotypes with different mechanisms of dormancy to determine the influence of environment on dormancy. ... Genotypes with embryo dormancy were consistently the most preharvest sprouting tolerant, even though this dormancy was influenced by the environmental conditions in the different seasons. Pyramiding the embryo component with the specific seed coat component and/or awnless head trait removed some of the environmental variation in preharvest sprouting tolerance, but this was generally considered excessive to the environmental requirements. The methods developed here, of field imposed stresses may provide a valuable tool to further understand the influence of environment on the regulation of dormancy, as different phenotypes can be made with the same genotype. Moisture stress, sudden changes in water supply or high temperatures during the late dough stages influenced dormancy phenotype and should be considered and avoided if possible when selecting locations and running trials for screening for genetic differences in preharvest sprouting tolerance. In the Western Australian context, the embryo component of dormancy appeared to be sufficient and should be adopted as the most important trait for breeding for preharvest sprouting tolerance.

Doctor of Philosophy / Department of Agronomy / Gina Brown-Guedira / Allan K. Fritz / Aegilops tauschii, the D-genome donor of modern wheat, has served as an important source of genetic variation in wheat breeding. The objective of this research was to characterize and utilize multiple Ae. tauschii-derived pest resistance genes contained in the germplasm KS96WGRC40. Two Ae. tauschii-derived genes, H23 and Cmc4, provide resistance to the Hessian fly (HF) and wheat curl mite (WCM), respectively. A linkage analysis of a testcross population estimated 32.67% recombination between H23 and Cmc4 on chromosome 6DS in wheat indicating that the two genes are not tightly linked as previous mapping reports show. Haplotype data of recombinant lines and physical mapping of linked microsatellite markers located Cmc4 distal to H23. Haplotype data indicated that both KS89WGRC04 and KS96WGRC40 have the distal portion of 6DS derived from Ae. tauschii. Microsatellite primer pairs BARC183 and GDM036 were more useful than the previously published linked markers in identifying lines carrying Cmc4 and H23, respectively. Through phenotypic selection and advancement within the testcross population, three TC1F2:4 lines were identified as homozygous resistant for H23 and Cmc4 and have the complete terminal segment of 6DS from Ae. tauschii. Two lines are more desirable than the original germplasm releases and can serve as a source of resistance to both HF and WCM in an elite background. A linkage analysis of a segregating recombinant inbred line population identified an Ae. tauschii-derived gene of major effect conferring resistance to Septoria leaf blotch (STB) and another Ae. tauschii-derived gene of major effect conferring resistance to soil-borne wheat mosaic virus (SBWMV) in the germplasm KS96WGRC40. The STB resistance gene in KS96WGRC40 is located in the distal 40% of the short arm of chromosome 7D flanked by microsatellite markers Xgwm044 and Xbarc126. Two previously reported STB genes, Stb4 and Stb5, have been mapped to 7DS in the same region as the STB resistance gene in KS96WGRC40. The uniqueness of the STB resistance genes on 7DS is questionable. The SBWMV resistance gene in KS96WGRC40 is located on chromosome 5DL linked to microsatellite marker Xcfd010. The SBWMV resistance gene within KS96WGRC40 was derived from TA2397 via KS95WGRC33.

T. aestivum ssp. spelta Iranian type has long been thought to potentially be the direct non-free threshing hexaploid progenitor. I evaluated a RIL population derived from a cross between CS and Iranian spelta accession P503 to identify loci suppressing free-threshabilty in P503. Identification of QTL associated with threshability in region known to harbor the Tg2A gene, and an inactive tg2D allele supported the hypothesis of Iranian spelta being derived from a more recent hybridization between free-threshing hexaploid and emmer wheat. Parastagonospora nodorum is an important fungal pathogen and secretes necrotrophic effectors that evoke cell death. In this research, a DH population segregating for Snn5 was used to saturate Snn5 region of chromosome 4B with molecular markers. The physical distance between Snn5 flanking markers was narrowed to 1.38 Mb with genetic distance of 2.8 cM. The markers developed in this study will provide a strong foundation for map-based cloning of Snn5.