Thesis (Ph. D.)--West Virginia University, 2007. / Title from document title page. Document formatted into pages; contains xi, 127 p. : ill. Includes abstract. Includes bibliographical references (p. 109-114).

Haploid plants are potentially valuable for the breeding of many crops particularly those with long reproductive cycles. In this investigation an attempt was made to determine the requirements of androgenesis of peach and cherry, a technique which has been used to produce large numbers of haploids in other species. A procedure used successfully with tobacco was verified. A preliminary survey of apple, cranberry and rose showed that they do not have the same requirements as tobacco. In most experiments with cherry and peach immature pollen was used. However the lack of differential responces of pollen to treatment made interpetation difficult. Hence responces of anthers and the growth of calli from somatic flower parts was used to assess different treatments instead of pollen growth. Calli emerged from cherry anthers when anthers were cultured on Nitsch's medium containing naphthalene acetic acid (NAA) and coconut milk and incubated in the dark at 30°C. Because these calli contained both diploid and tetraploid cells and because of the lack of abnormal pollen development in the anthers it was concluded that the calli probably originated from anther connective tissue rather than pollen. In experiments with mature cherry pollen several different types of abnormal pollen growth were observed including multicellular pollen grains. A comparison of the requirements for species in which androgenesis has been demonstrated is discussed. In some species components making up the requirements appear to form a pattern which if verified by reports from other species will be valuable in future investigations. Suggestions are made as to the direction which further investigations of the requirements for androgenesis should take. / Land and Food Systems, Faculty of / Graduate

The production of barley (Hordeum vulgare L.) haploids by crossing with H. bulbosum is a widely used tool in breeding and genetics. Certain barley genotypes have low seed set in this interspecific cross, a phenomenon known as partial incompatibility. Haploid production efficiency and gamete sampling are important issues with the bulbosum technique, particularly when partially incompatible genotypes are used. An in vitro floret culture system was developed that substantially increases haploid production efficiency by optimizing caryopsis growth, haploid embryo development, and plant regeneration. The individual and combined effects of three plant growth regulators (2,4-D, GA₃ and kinetin) on haploid production efficiency and its determinants were compared in the floret culture system. 2,4-D alone was superior to GA₃ alone in haploid production efficiency. 2,4-D alone or kinetin + 2,4-D are recommended for the purpose of haploid production in floret culture using the bulbosum method. Partial incompatibility between H. vulgare and H. bulbosum was studied by doubled haploid progeny analysis. Two different loci were hypothesized to account for the inheritance of partial incompatibility in the crosses of Vada x Klages, Harrington x Klages, and Vada x Harrington. The partial incompatibility gene in Harrington was found to be recessive. The dominant nature of the partial incompatibility gene (Inc) in Vada was confirmed. An association between the (Inc) gene and a deficiency in a stigma/stylodium-specific high pl protein was found in the cosegregation analysis of doubled haploid progeny. The Inc gene may be linked to the gene coding for the stigma/stylodium-specific protein, or the Inc gene may regulate expression of the protein-encoding locus. Segregation analysis of Mendelian markers in doubled haploid progeny showed that there is no evidence that the partial incompatibility status of the parents has an effect on gamete sampling by the bulbosum technique. / Graduation date: 1991

Thesis (Ph.D.) - University of Queensland, 2005. / Includes bibliography.

No description available.

A dissertation submitted to the Faculty of Science, University of the Witwatersrand, in fulfillment of the requirements for the degree of Master of Science Johannesburg, August 2012 / It is known that infectious agents elicit different responses in different individuals which strengthens the view that susceptibility and resistance to infectious diseases has a genetic component. These differences in susceptibility to disease can be observed in populations. APOBEC3G is a member of the cytidine deaminase gene family located on chromosome 22. It is crucial in non-permissive cells as it functions as part of the innate immunity system and is an inhibitor of the HIV-1 accessory protein vif. The goal of the study was to develop genotyping assays and estimate allele frequencies. Thus, genetic variation within APOBEC3G was identified and characterized in black South Africans. Indirect genotyping assays were designed to amplify regions within the upstream non-coding region, and in exon 4 of the coding region of the gene. Selected polymorphisms were then genotyped using allele-specific PCR, RFLP-PCR and Pyrosequencing™ assays. Reanalysis of sequence data from 2003 showed numerous SNPs were well represented. Comparison of sequence data at various SNPs showed that allele frequencies were similar to frequencies in other African populations. The only sequenced SNP that deviated from the frequencies in Ensembl was -590. Thus the sequencing was a useful tool for detection of variation. ASA proved to be the least reliable genotyping technique as the minor allele frequency of -571 (0.59) deviated from the published frequency of 0.894 in Africans. RFLP analysis proved more reliable for genotyping -571 and H186R. The minor allele frequency was estimated to be 0.84 and 0.32 for -571 and H186R respectively. The frequency of H186R is similar to published data from An et al (2004) and Reddy et al (2010). If SNPs are in LD they occur together on the same haplotype more often than by chance. Usually SNPs that are in LD are in close proximity. However our data suggests -571 and H186R SNPs which are 5kb apart are not in LD. A LD map of chromosome 22 shows highly variable pattern of LD (Dawson et al, 2002). Widespread regions of nearly complete LD up to 804 kb in length are intermingled with regions of little or uundetectable LD. Haplotype analysis showed the most frequent haplotype was GA. This was the most frequent haplotype when the sample types were subdivided according to spoken language. in comparison to studies from An et al, (2004) D’ of the two SNPs was estimated at 0.967. The linkage disequilibrium (LD) revealed a non-independence of allele segregation because the loci analyzed were strongly linked in the Apobec 3 G gene. The data are consistent with greater genetic diversity of African populations and can form the basis for further evaluation of the role of variation in this gene in response to HIV.

Dissertation (Ph.D.)--University of Toledo, 2009. / "In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biomedical Science." Title from title page of PDF document. Bibliography: p. 87-96.

Maintenance of genomic integrity is essential for the survival of an organism and its ability to pass genetic information to its progeny. However, DNA is constantly exposed to exogenous and endogenous sources of damage, which demands cells to possess DNA repair mechanisms. Of the many forms of DNA damage, double-strand breaks (DSBs) are particularly cytotoxic DNA lesions that cause genome instability and cell lethality, but also provide opportunities to manipulate the genome via repair. One of the major DSB repair pathways shared between single-celled yeast and humans is homologous recombination (HR). HR is initiated by the evolutionarily conserved Mre11-Rad50-Xrs2/Nbs1 (MRX in yeast, MRN in mammals) complex. The MRX complex has a multitude of functions such as damage sensing, adduct removal from DSB ends, and end tethering – a process to maintain the two ends of a DSB in close proximity. The role of the MRX complex has been uncovered by studying the repair of DSBs generated from meganucleases such as HO and I-SceI. However, it is unclear if this knowledge translates to the repair of DSBs from genome editing nucleases such as TALEN and CRISPR-Cas9 (Cas9), as these nucleases create DSBs with different end polarities. While the repair efficiencies and outcomes of TALEN and Cas9 are actively studied, less is known about the earlier stages of repair. The objective of this thesis is to examine the role of the MRX complex in repair processes at both ends of a DSB after cleavage with I-SceI, TALEN, and Cas9 in vivo using the model organism Saccharomyces cerevisiae. In Chapter 1, I describe the importance of DSB repair, a summary of HR and its sub-pathways, the functions of the MRX complex, and properties of I-SceI, TALEN, and Cas9. The materials and methods used in this thesis are detailed in Chapter 2. The work described in Chapter 3 focuses on end tethering and recruitment of downstream repair proteins in haploid cells. I find that DSB ends from the three nucleases all depend on the MRX complex for end tethering, and that initial end polarity does not affect tethering. DSBs created by Cas9 show greater dependence on the Mre11 nuclease of the MRX complex for Rad52 recruitment compared to DSBs from I-SceI and TALEN. Despite Mre11-dependent end processing and Rad52 recruitment at Cas9-induced DSBs, Cas9 stays bound to one DNA end after cleavage, irrespective of the MRX complex. These results suggest that Mre11 exonuclease activity required for adduct removal from DSB ends is not critical for Rad52 recruitment, and that Mre11 endonuclease activity may be driving processing of Cas9-bound DSBs. I also find that MRX tethers DSB ends even after Rad52 recruitment, and unexpectedly, untethered ends are processed asymmetrically in the absence of MRX for all three nucleases. In Chapter 4, I explore the interaction of DSB ends with their repair template, the intact homologous chromosome, in diploid cells. The primary goal is to monitor interhomolog contact in real time from homology search to completion of HR. Although technical limitations make it difficult to capture the entire HR program from DSB formation to repair, I show that untethered ends interact with the homolog separately in the absence of the MRX complex. Similar to haploids, diploid cells display defects in end tethering and end processing without the MRX complex. Repair outcomes of WT cells show an even distribution of G2 crossovers and non-crossovers, while pre-replication crossovers and break-induced replication are undetected. Overall, the results in this thesis provide insight into the functions of the MRX complex in repairing different DSB ends created by I-SceI, TALEN, and Cas9. In Chapter 5, I summarize all of these findings and discuss the motivation for future cell biology studies of HR.

Doubled haploidy (DH) is a plant breeding technique that is often utilized by plant breeders to minimize the time required to reach homozygosity in breeding lines. The first objective of this study was to compare two methods of generating DH lines in tobacco (Nicotiana tabacum L.). Inbred burley tobacco varieties TN 90LC and GR 149LC were used to produce both androgenic derived doubled haploids (ADDH) and maternally derived doubled haploids (MDDH). The relative agronomic performance of TN 90LC and GR 149 LC ADDH and MDDH lines was compared when used either as pure-line cultivars or when used for the production of the KT 204LC and TN 97LC hybrid cultivars, respectively. The ADDH method was more efficient than the MDDH method in generating large numbers of haploid plants. On average the ADDH TN 90LC population was statistically inferior to the inbred TN 90LC for several agronomic traits; this inferiority of the ADDH method was not observed in the GR 149LC populations. For both genotypes, the MDDH populations were comparable to the inbred parental genotypes. The ADDH method was inferior for TN 90LC, but several individual TN 90LC ADDH lines were equal or superior to the inbred source. The agronomic variability observed in both ADDH and MDDH lines was decreased when they were used to produce hybrid cultivars. Less variation was observed in the DH-derived hybrids KT 204LC and TN 97LC compared to the ADDH and MDDH TN90LC and GR149LC parental lines, respectively. The significant inferiority of ADDH TN 90 lines in comparison to inbred TN 90LC was not observed in the ADDH derived KT 204 population compared to KT 204LC. The second objective of this study was to compare DH Lines derived from an F1 breeding population versus DH lines derived from a segregating F2 population where plants used for DH were pre-screened for quantitatively inherited resistance to soil-borne diseases black shank (Phytophthora nicotianae) and/or Fusarium wilt (Fusarium oxysporum f. nicotianae). There was a clear difference in susceptibility to black shank between the F1 and F2 derived DH populations, both in terms of average disease incidence, and more importantly, in the percentage of individual lines displaying high disease resistance. For two different burley crosses, DH lines derived from the F1 generation were considerably more susceptible to black shank than DH lines derived from the F2 generation. No differences in the incidence of Fusarium wilt were observed between DH lines of F1 and F2 generations; this was likely due to low overall disease incidence. Although delaying the DH process in tobacco from the F1 to the F2 generation could add time to the development of homozygous breeding lines, the delay may be offset by having to screen fewer finished DH lines to identify superior lines.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.