Sugarcane is an economically important crop and its high demand has necessitated the use of biotechnology methods to produce and accelerate the production of desirable genotypes. One such method is genetic transformation. However, as sugarcane is a highly polyploid crop, which originated from interspecific crosses between Saccharum spontaneum and S. officinarum, efforts to transform it are inhibited by transgene promoter silencing. As ancestral lines have a simpler genetic makeup than modern varieties, they may be useful to test promoter function. Intrinsic to the generation of transgenic plants is the ability to produce plants from specific species and varieties, for which an indirect method of regeneration is needed. Consequently, the first objective of this study was to determine a high yielding protocol for somatic embryogenic calli. The second was to transform such calli and produce regenerated plants to assess transgene expression. A preliminary study was conducted using eight ancestral varieties to determine which were the most responsive in culture. Leaf roll disks were cultured on 5 mg.1¯¹ 2, 4-D and callus production was assessed. Based on these results and the availability of plant material, S. spontaneum Nigeria 1, S. spontaneum Nigeria 2, S. spontaneum Coimbatore, S. officinarum NG 77-69, and S. officinarum Black Cheribon and the commercial polyploid variety NCo376 were selected and tested on 11 different callus induction media. The S. spontaneum variety that generated the highest percentage of leaf disks that produced callus and plant yield was Nigeria 1 (61 % and 259 plants/10 disks, respectively), whilst the S. officinarum variety was Black Cheribon (75 % and 90 plants/10 disks, respectively). The best media for both comprised of MS salts and vitamins, 20 g.1¯¹ sucrose, 0.5 g.1¯¹ casein hydrolysate 5 mg.1¯¹ 2, 4-D and 8 g.1¯¹ agar. NCo376 produced the most amount of callus (93 %) when cultured on media containing 3 mg.1¯¹ 2, 4-D and gave a final yield of 450 plants/10 disks. Based on the yields obtained above and the availability of plant material, the varieties S. spontaneum Nigeria 1 and S. officinarum NG77-69 were selected for genetic transformation studies. Calli of these varieties as well as that of NCo376 were microprojectile bombarded with either pEmuKN + pAHC27 or pEmuKN + pR₁₁F¯. Following bombardment, the calli were cultured onto paromomycin-containing (1 ml.1¯¹) selection media and regenerated plants were obtained after 8-12 weeks. Transgene integration into the plant genome was assessed using PCR and qPCR techniques, and indicated that all NCo376 plantlets contained the GUS and npt II transgenes. However, only 4 out of 5 and 2 out of 3 S. officinarum NG77-69 plants transformed with pAHC27 and pR₁₁F¯- respectively, and 6 out of 10 S. spontaneum Nigeria 1 plants transformed with pR₁₁F¯- contained these transgenes. The transformation efficiencies achieved for NCo376, for the constructs pAHC27 and pR₁₁F¯- was 0.27 and 0.33 transgenic plants/blast, respectively. For NG77-69 it was 0.27 and 0.13 transgenic plants/blast, whilst that of Nigeria 1 was 0.20 and 0.40 transgenic plants/blast. Stable transgene expression in acclimatized plants was then assessed using a histochemical GUS assay and none of the plants expressed the GUS gene. / Thesis (M.Sc.)-University of KwaZulu-Natal, Westville, 2013.

Mapping studies of the centromeric region of the human Y chromosome Construction of a map of a human centromeric region is very important in order to understand the organisation of this essential part of the chromosome. A YAC contig map has been assembled of the pericentric 10 Mb of the human Y chromosome, giving coverage of Yp from the large X-Y homologous region through to the alphoid satellite of the centromere, and from the alphoid DNA to the proximal unique sequences on Yq. The Yp map has one remaining gap between TSPY1 and the AMELY region, while two gaps separate the satellite region on Yq from the other two contigs. After constructing the map, the known genes were localised to the region. One Yq gene, DFFRY, was discounted as a potential anti-Turner syndrome gene by analysis of rearranged Y chromosomes. Detection of a block of duplicated sequence on Yp led to the confirmation of the existence of an inversion polymorphism, which was then found to be correlated with a major subclass of sex-reversed individuals, who have X-Y chromosomal breakpoints within the inverted region. These results not only give a far more extensive and detailed map of this region than before, but also show that understanding the organisation of the region has important consequences for a number of genetic disorders.

A modification of bulked segregant analysis was used to raise the density of markers in a 34.5 cM region between Ugp2 and Ugp1, on chromosome 3 of the Harrington x TR306 barley Hordeum vulgare L.) cross. A computer program was used to select pools contrasting for parental alleles at the target site. Of 257 RAPD primers tested on DNA pools, one, UBC 508, detected a polymorphic DNA fragment (UBC508(C)). It mapped 10.2 cM distal to Ugp2. Two additional DNA polymorphisms, (UBC508(A) and UBC508(B)), mapped on chromosome 2. An additional marker, BCD 1796B, mapped 4.9 cM proximal to Ugp1. Both strands of the UBC508(C) fragment were sequenced. They were 588 bp long and had some homology to a region of the DNA that regulates transcription of the H. vulgare pazx gene encoding protein Zx. The effect of adding new marker(s) on the QTL analysis of agronomic and quality traits of barley, was investigated. For extract beta-glucan, a new peak was identified in the analysis when only UBC508(C) or when both UBC508(C) and BCD1796B were added. For fine-coarse difference a QTL x E interaction peak was detected when only BCD 1796B was added or when both UBC508(C) and BCD 1796B were added.

This thesis reports the results of genetic and molecular analysis of polytene region 38 of D. melanogaster. / A detailed genetic map of the region has been constructed through the complementation analysis of 22 genes with 37 deficiency stocks, providing 48 breakpoints. 44 of the breakpoints have been precisely mapped and separate the complementation groups into 16 distinct intervals. / Molecular characterisation was achieved using a combination of computational sequence analysis and experimental techniques. In this manner, 46 new transcription units and their developmental expression patterns were identified and the physical location of 7 deficiency breakpoints revealed. Links have been established between the genetic and the physical maps using STSs and sequence information from previously cloned genes. This allowed the possible correlation of 4 previously genetically defined genes to their transcription units.

Two studies were conducted on gene mapping analysis. For the first study, genetic simulation experiments were conducted to address the effects of marker density, method of mapping analysis, and gaps in a marker map on the efficiency of QTL detection and the accuracy of QTL parameter estimation. The simulated genome consisted of seven chromosomes with seven or eight segregating QTL affecting the simulated quantitative trait. A set of six randomly segregating QTL outside the test region was consistently used to represent 40% of phenotypic variation. An individual QTL or a linkage block of two QTL on a target chromosome contributed 10% of phenotypic variation. The marker map was either dense (with markers every 4 cM) or sparse (with markers every 20 cM). The gap in the marker map was either 32 cM or 56 cM. Interval mapping and composite interval mapping were used to map QTL on the target chromosome. A dense map provided more power of QTL detection, better accuracy of QTL parameter estimation, and higher false-positive error rates for the target chromosome than a sparse map. Composite interval mapping provided more power of QTL detection, better accuracy of QTL parameter estimation, and lower false-positive error rates than interval mapping. Presence of a large gap in a marker map affected QTL detection and QTL parameter estimation for a QTL inside or near the gap. The use of a dense map with composite interval mapping was the most efficient combination tested in this study. For the second study, a mixed factorial regression model for interval mapping was developed for conducting QTL-by-environment interaction analysis and for providing inferences about QTL that are applicable beyond the environments used in the experiments. Genetic simulation was used to test the model for the power of detecting QTL-by-environment interaction and identifying the types of such interaction as crossover or non-crossover, and for the accuracy of estimating QTL parameters. The model prov

Thesis (Ph.D.)-- University of Adelaide, Dept. of Plant Science, 2001. / Includes bibliographical references (leaves 192-211).

Thesis (Ph. D.)--Illinois State University, 1996. / Title from title page screen, viewed May 31, 2006. Dissertation Committee: David F. Weber (chair), Alan J. Katz, Marjorie A. Jones, Radheshyam K. Jayaswal, Jefferey A. Dole. Includes bibliographical references (leaves 96-108) and abstract. Also available in print.

M.Sc. (Biochemistry) / A 14-member, closely-spaced cluster of genes coding for leucine-rich repeat receptor-like kinases (LRR-RLKs) is located on chromosome 1 of Arabidopsis thaliana. Following on from previous microarray studies that found some of the members of this cluster to be upregulated in response to biotic stressors, including the bacterial elicitor flg22, the present study sought to confirm, using a luciferase-based protoplast assay, that flg22 does in fact induce the expression of the genes, and then to investigate the promoters of the genes. The promoters of At1g51790, At1g51850 and At1g51890 responded positively in this particular assay, and bioinformatic analyses determined that W-boxes are over-represented in the cloned regions. Mutational inactivation of individual W-boxes in the promoter of At1g51790 drastically reduced the flg22 response, except for the W-box closest to the start site, which seemed to increase both basal and flg22-inducible expression. In the promoter of At1g51850, mutational inactivation of either or both of its W-box dyads resulted in virtually no flg22 inducibility. The deletion of 6 W-boxes in the promoter of At1g51890, done via truncation, drastically reduced both its basal expression and its inducible response to flg22. These results provide evidence that W-box cis-elements are responsible for the upregulation of these LRR-RLKs in response to flg22. WRKYs -7, -11, -22,and -26 were found bioinformatically to have similar expression patterns to some of the genes in the cluster, and are thus good candidates to investigate as transcriptional regulators of the cluster in future studies.

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