Thesis (Ph. D.)--Texas A&M University, 2008. / "Major Subject: Physiology of Reproduction" Title from author supplied metadata (automated record created on Oct. 13, 2008.) Vita. Abstract. Includes bibliographical references.

Thesis (MScAgric)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: Reproductive bud development is the first step of sexual reproduction of plants and is a major factor determining yield. For consistent yields in pears, knowledge of the development of the reproductive bud is required. The development of the reproductive bud with emphasis on the primordial bourse shoot was studied. The bourse shoot is important because of the ability of a bourse shoot to become reproductive, which can lead to bourse-overbourse bearing. The progression of reproductive bud development in 'Forelle' and 'Rosemarie' was studied from primordial bourse shoot initiation until dormancy the following season. The primordial bourse shoot was initiated in January 2003. The primordial bourse shoot progressed during dormancy with a plastochron length of 60 days in June. The rate of preformed leaf formation increased rapidly until August when the plastochron was 5 days. 'Rosemarie's' primordial bourse shoot development was more advanced at full bloom, which is a possible reason for the higher bourse-aver-bourse bearing habit of the cultivar. The number of leaves of the bourse shoot was more than with the 'Forelle', which has a low tendency for bourseover- bourse bearing. Flower initiation of the terminal bourse shoot bud was 56 and 77 d.a.f.b. for the 'Forelle' and 'Rosemarie', respectively. Initiation was well correlated with bourse shoot growth cessation. An increase in mitotic activity occurred during flower differentiation of the terminal bourse bud with a peak of floral appendage formation during December and January. From February until dormancy enlargement of the floral parts took place. The influence of severe dormant pruning and the quality of 2- year-old wood, on reproductive bud sink strength of 'Packham's Triumph' trees were investigated. The sink strength of primary growth increased by 40 % for short bearing units (SBU's) and 140 % for thick bearing units (BU's) as compared to long bearing units (LBU's) and thin BU's respectively. The increase in primary growth was due to better fruit set and larger fruit in the SBU's and the thick BU's. Since the increase in primary growth is less for SBU's compared to thick BU's the conclusion, which can be made, is that branch diameter influenced sink strength more than branch length. With increase in branch diameter the size of the xylem transport system increases more than that of the phloem. The increase in xylem transported metabolites, in particular root derived cytokinin seems to predominantly influence the magnitude of the sink strength. The influence of scoring and 6-benzyladenine (BA) application, during the flower induction phase on 'Doyenne du Cornice' and 'Rosemarie', were studied. Scoring caused a 50 % increase in fruit number of 'Doyenne du Cornice', which resulted in a 38 % increase in yield compared to the control. The increase in fruit number was due to higher percentage reproductive buds and improved bud quality. For the 'Rosemarie' scoring at the correct time resulted in a 40 % increase in reproductive buds. Scoring disrupts basipetal transport in the phloem, which results in the removal of apical dominance and an increase in root derived cytokinin's. More meristems can respond to inductive conditions and high quality reproductive buds develop. The combination of BA and scoring, lead to more flowers per inflorescence in 'Doyenne du Cornice'. / AFRIKAANSE OPSOMMING: Reproduktiewe knopontwikkeling by die peer Reproduktiewe knopontwikkeling is die eerste fase van reproduktiewe voortplanting in plante en is belangrik, want dit bepaal die oesgrootte. Vir konstante oeste in peerproduksie moet reproduktiewe knopontwikkeling verstaan word. Die ontwikkeling van die reproduktiewe knop, meer spesifiek die primordiale beursloot is bestudeer. Die beursloot is belangrik a.g.v. die beurs-oorbeurs drawyse. Die ontwikkeling van die reproduktiewe knop van 'Forelle' en 'Rosemarie' is gevolg. Die primordiale beursloot is Januarie 2003 geinisieër en ontwikkel tydens die boom se dormante fase met 'n plastochron van 60 dae gedurende Junie. Preformeerde blaarvormingstempo neem toe tot Augustus waar die plastochron 5 dae is. 'Rosemarie' se primordiale beursloot ontwikkeling was meer gevorderd teen volblom, met meer beurslootblare reeds teenwoordig. Dit is dalk die rede vir die hoër tendens van beurs-oor-beurs drag van die kultivar in vergelyking met 'Forelle'. Die terminale beurslootknop is 56 en 77 dae na volblom geïniseer vir 'Forelle' en 'Rosemarie' respektiewelik. Inisiasie was goed gekorreleerd met beeïndiging van beurslootgroei. Blomdifferensiasie het inisiasie gevolg met 'n toename in mitotiese aktiwiteit, blomaanhangsel vorming het 'n piek bereik III Desember en Januarie. Vanaf Februarie tot dormansie het ontwikkeling hoofsaaklik in vergroting van blomorgane plaasgevind. Die invloed van strawwe dormante snoei en die kwaliteit van 2-jaar-oue dra-eenhede op sinksterkte van reproduktiewe knoppe van 'Packham's Triumph' pere is ondersoek tydens die 2002/03 seisoen. Die sinksterkte van primêre groei het 40 % toegeneem vir kort dra-eenhede en 140 % vir dik dra-eenhede in vergelyking met lang dra-eenhede en dun dra-eenhede respektiewelik. Die toename in primêre groei vir kort en dik dra-eenhede kon toegeskryf word aan beter vrugset en groter vrugte. Omdat die toename in sinksterkte minder is vir kort dra-eenhede in vergelyking met dik dra-eenhede kan die afleiding gemaak word dat dra-eenheiddikte sinksterkte meer beïnvloed as dra-eenheidlengte. Met 'n toename in dra-eenheiddikte, neem xileemvaatweefsel meer toe as floeëmvaatweefsel. 'n Toename in xileem getranslokeerde metaboliete, meer spesifiek wortel vervaardigde sitokiniene beïnvloed die sinksterkte van die reproduktiewe knoppe. Die invloed van ringelering (ringsnit deur floeëm sonder bas verwydering) en 6-bensielaldenien (BA) toediening tydens reproduktiewe knopinduksie van 'Doyenne du Cornice' en 'Rosemarie' is ondersoek. Ringelering het 'n 50 % toename in vrugaantal veroorsaak wat die oes met 38 % laat toeneem het in vergelyking met die kontrole, vir 'Doyenne du Cornice'. Die toename in vrugte was a.g.v. 'n hoër persentasie reproduktiewe knoppe en toename in blomkwaliteit. By 'Rosemarie' het ringelering 'n 40 % toename in reproduktiewe knoppe bewerkstellig. Ringelering onderbreek die basipetale vervoer in die floeëm, verwyder dus apikale dominansie, met die gevolg dat daar 'n toename in wortel geproduseerde sitokiniene is. Meer meristeme reageer op induktiewe toestande en reproduktiewe knoppe van hoë kwaliteit vorm. Die kombinasie van ringelering en BA -toediening het meer blomme per reproduktiewe knop veroorsaak.

Thesis (MScAgric)--University of Stellenbosch, 2000. / ENGLISH ABSTRACT: Ineffective pollination of the main cultivar with the pollinator cultivar is due to either an incompatibility problem between the main and pollinator cultivar, or because the flowering times of the main and pollinator cultivars do not overlap adequately. Three trials were conducted to try and find a more effective way to determine cultivar compatibility and to group cultivars together according to their budburst reaction to chilling. Most apple cultivars are self-incompatible and need cross-pollination for fruit set. Due to differences in the genetically defined fertilisation compatibility between the pollen from the male parent (pollinator) and the egg cell of the female parent, various apple pollinators differ in their ability to set fruit with viable seed. Fruit weight and size are positively correlated with seed set although it has been found that the pollinator can have a direct influence on fruit quality. 'African Red' apple trees on M7 rootstock in an evaluation block on a commercial farm in the Koue Bokkeveld region (32°55'N 19°27'E, Mediterranean climate, ;::::1060Utah chill units, and ;::::530mm rainfall annually; altitude 966 m) were used to assess the influence of 5 pollinators ('Granny Smith', 'Winter Banana', 'Cripps' Pink', 'Cripps' Red' and 'Simpson Crab') on fruit set, fruit weight and length and diameter. The degree to which 'African Red' is self-compatible was also assessed and the effect of flowering position ("king" vs. lateral) on fruit quality was determined. None of the pollinators showed a significantly higher fruit set. No differences in fruit set were found between the "king" and lateral flowering positions. No significant differences were found in the average number or weight of well developed seeds between pollinators. In both years fruit weight was significantly correlated to seed number for all five pollinator cultivars. In 1998 'Simpson crab' gave fruit that were significantly more elongated than those of 'Cripps' Pink'. 'African Red' is highly self incompatible. Compatibility assessments that are based on the number of fruit that develop after the flowers ofthe main cultivar had been hand pollinated in field trials are a time-consuming process. Allele-specific PCR amplification for some of the known S-alleles of the incompatibility S-gene (S2, S3, S5, S7 and S9) was carried out to successfully predict the compatibility of genotypes. The results compared well with that found in literature. For all the Malus domestica cultivars tested at least one, but in some instances both alleles of the S-gene were determined. 'Simpson crab' (Malus baccata) did, however, not possess any of the tested S-alleles. One-year-old, ca. 40 mm long shoots of various apple cultivars were selected from commercial orchards in both the Elgin [34°S, 305 m, ca. 750 chill units (CU) (Richardson et al., 1974)] and Koue Bokkeveld (33°S, 945 m, ca. 1300 CU) regions of the Western Cape, South Africa in two consecutive years (1998 and 1999). Shoots were forced at a constant 25°C with continuous illumination after receiving their allocated chill units. The effect of chilling period on the budburst of each cultivar in both regions was estimated by determining, 1) the total proportion of budburst (%Bb), 2) the proportion of shoots with terminal budburst (%TBb), and 3) the rate of budburst [lI(days to 25% budburst)]. It was found that these indices differed significantly between cultivars, and within cultivars between areas, as far as budburst patterns, in reaction to chilling, were concerned. The rate of budburst was the most consistent in describing the reaction of buds to different chilling periods and could be used to group cultivars together according to their budburst reaction to chilling. / AFRIKAANSE OPSOMMING: KRUISBESTUIWINGSBIOLOGIE VAN APPELS MET SPESIALE VERWYSING NA 'AFRICAN RED'. Oneffektiewe kruisbestuiwing III die boord kan toegeskryf word aan Of onverenigbaarheid tussen die hoof- en bestuiwingskultivar Of as gevolg van die blomtyd van die kruisbestuiwer wat nie genoegsaam oorvleuel met die van die hoofkultivar nie. Drie proewe is uitgevoer om 'n meer effektiewe proses daar te stel vir die toets van kultivarverenigbaarheid en om kultivars te probeer groepeer na gelang van hul reaksie op bepaalde hoeveelhede koue. Meeste appelkultivars is selfonverenigbaar en benodig kruisbestuiwing vir genoegsame vrugset. As gevolg van verskille III die geneties gedefinieerde bevrugtingsverenigbaarheid tussen die stuifmeel van die manlike ouer (bestuiwer) en die eiersel van die vroulike ouer (hoofkultivar), verskil bestuiwers in hul vermoë om vrugte met sade te set. Vruggrootte en -massa is positief gekorreleerd met saadset alhoewel dit al gevind is dat die bestuiwer op sig self ook 'n invloed op vrugkwaliteit kan hê. 'African Red' appelbome op M7 onderstamme, in 'n evaluasie blok op 'n kommersiële plaas in die Koue Bokkeveld (32°55'N 19°27'E, Meditereense klimaat, ::::1060 Utah koue eenhede, en ::::530mmjaarlikse reënval; ligging 966 m), is gebruik om die invloed van 5 bestuiwers ('Granny Smith', 'Winter Banana', 'Cripps' Pink', 'Cripps' Red' and 'Simpson Crab') op vrugset, vrugmassa, -lengte en -deursnee oor twee seisoene te bepaal. Die mate waartoe 'African Red' self onverenigbaar is en die effek van blomposisie ("king" vs laterale blom) op vrugkwaliteit is ook bepaal. Geen een van die bestuiwers het vrugset beduidend beïnvloed nie. Ook is daar geen verskille gevind tussen die "king" en laterale blomposisies t.o.v. vrugset nie. Geen beduidende verskille is tussen bestuiwers gevind in die gemiddelde aantal of gewig van volsade geset nie .. In albei jare was die vrugmassa beduidend gekorreleerd met saadset vir al vyf bestuiwerkultivars. In 1998 het 'Simpson Crab' vrugte geset wat beduidend langer was as vrugte wat geset het toe 'Cripps' Pink' as bestuiwer gebruik is. Daar is ook gevind dat 'African Red' hoogs selfonverenigbaar is. Verenigbaarheidstoetse wat gebaseer is op die aantal vruggles wat ontwikkel nadat blomme van die hoofkultivar met die hand bestuif is, is 'n tydsame proses. Allele spesifieke PCR amplifikasie vir bekende S-allele van die onverenigbaarheids S-geen (S2, S3, S5 S7en S9) is suksesvol uitgevoer om die verenigbaarheid van genotipes vooraf te bepaal. Die resultate het goed vergelyk met wat in literatuur gevind is. Vir al die Malus domestica spesies wat getoets is, is ten minste een, en in sommige gevalle twee, van die S-allele gevind. Die blomappel 'Simpson' (Malus baccata) het egter nie een van die vyf S-allele opgelewer nie Een-jaar-oue, 40 mm lang lote van verskeie appelkultivars, is in twee opeenvolgende jare (1998 en 1999) vanuit kommersiële boorde in beide die Elgin [34°S, 305 m, ca. 750 koue eenhede (CU) (Richardson et al., 1974)] and Koue Bokkeveld (33°S, 945 m, ca. 1300 CU) areas van die Wes Kaap gsny. Die lote is geforseer om te bot by 'n konstante temperatuur van 25°C met deurlopende beligting, nadat elke groep lote aan 'n bepaalde hoeveelheid koue blootgestel is. Die effek van koue op bot van elke kultivar in beide areas is bepaal deur, 1) die totale persentasie knoppe wat gebot het, 2) die persentasie terminale knoppe wat gebot het, en 3) die tempo van bot [l/(dae tot 25% bot)] te meet. Daar is gevind dat bo-genoemde parameters beduidend tussen kultivars, en binne kultivars tussen areas, verskil. As 'n beskrywing van die reaksie van knoppe op koue het die tempo van bot die mees konstante resultate oor die twee opeenvolgende seisoene gelewer en kon hierdie parameter gebruik word om kultivars in groepe, na gelang van hul reaksie op koue, in te deel.

Luteinizing hormone (LH), one of the two gonadotropin hormones released from the pituitary gland, binds its receptor (LHR) in the gonads to initiate steroid hormone production, as well as gametogenesis and ovulation. Mutations of amino acid sequence within the receptor can render it either inactive or constitutively active. All activating mutations result in male-limited precocious puberty. Males afflicted with this condition undergo puberty around 4 years of age, with elevated testosterone levels and premature skeletal development. In order to better understand how chronic ligand-mediated activation of the LHR affects gonadal development and function, a mouse model expressing a yoked hormone-receptor (YHR) complex, engineered by covalently linking the hormone human chorionic gonadotropin to the rat LHR, has been studied. YHR+ males have prepubertally elevated testosterone and decreased gonadotropin levels. Histological evaluation of the testes of these animals show significantly smaller seminiferous tubules and Leydig cell clusters. Finally, testis gene expression analysis revealed a significant decrease in the relative mRNA expression of three Leydig cell specific genes. Based on these results, it was hypothesized that premature activation of the LHR impairs postnatal Leydig cell development. In the testis there are two morphologically and developmentally distinct populations of Leydig cells, the fetal and the adult. The first objective of this study was to quantify the populations of cells in the adult Leydig cell lineage in both the YHR+ and the WT controls. Real-time RT-PCR, for markers of the immature and adult Leydig cell populations, as well as Leydig cell quantification, suggested a delay in adult Leydig cell development. Interestingly, there was a significant increase in the fetal Leydig cell population in the YHR+ mice. The second objective was to determine if the decrease in the adult population is due to either a decrease in proliferation or an increase in apoptosis in the YHR+ animal. There was not a difference in apoptosis between the WT and the YHR+ at any age examined, however, there was a decrease in progenitor Leydig cell proliferation in the YHR+ animals at 2 weeks of age. The final objective was to determine if elevated neonatal testosterone levels impairs the development of the adult Leydig cell population. Seven-day old WT pups were subjected to testosterone supplementation via subdermal implant. Quantification of the total Leydig cell population revealed a significant decrease in the number of adult Leydig cells in the testosterone-treated group similar to that seen in the YHR+ animal. Taken together, these data suggest that elevated neonatal testosterone levels resulting from premature LHR activation inhibits the proliferation of progenitor Leydig cells, resulting in fewer adult Leydig cells in the YHR+ animals.

Luteinizing hormone (LH), one of the two gonadotropin hormones released from the anterior pituitary gland, binds to its receptor (LHR) in the gonads to stimulate steroid hormone production, in addition to ovulation and gametogenesis. Mutations of the receptors amino acid sequence have the ability to either constitutively activate or inactivate it. All activating mutations result in male-limited precocious puberty. Males with this condition undergo puberty around 4 years of age, and have a premature elevation in testosterone levels and premature skeletal development. In order to understand how chronic ligand-mediated activation of the LHR affects gonadal development and function, a mouse model expressing a yoked hormone-receptor (YHR) complex, engineered by covalently linking the hormone human chorionic gonadotropin to the rat LHR, has been studied. YHR+ males have prepubertally elevated testosterone and decreased gonadotropin levels, smaller testis, and smaller average seminiferous tubule diameters when compared to wild type (WT) animals. In a preliminary breeding study it was shown that YHR+ males were sub-fertile. Based the phenotype exhibited by the YHR+ mice, it was hypothesized that increased levels of testosterone in addition to decreased gonadotropin hormone levels in neonatal and prepubertal mice that results from premature activation of the luteinizing hormone receptor causes spermatogenesis to be impaired. The first objective of this study was to determine if there was a difference in the testicular germ cell and Sertoli cell populations in the WT and YHR+ animals using flow cytometry and systematic Sertoli cell counting, respectively. There was no difference in the Sertoli cell population between YHR+ animals and WT controls, but there were significantly fewer total germ cells in YHR+ animals at 10 days and from 4 weeks through adulthood. The second objective was to calculate the daily sperm production in the testis and epididymis of WT and YHR+ animals in order to determine if there is a further decrease in the total sperm count due to an epididymal dysfunction. Interestingly, there were significantly fewer sperm calculated in the caput/corpus region of the epididymis in YHR+ males at 12 weeks of age, but not in the testis and cauda epididymis. Furthermore, the daily sperm production in WT and YHR+ mice at 16 weeks of age were not significantly different. The final objective was to determine if the decrease in germ cells observed in YHR+ animals is the result of decreased proliferation or an increase in either germ cell or Sertoli cell apoptosis. Quantification of germ cell and Sertoli cell proliferation revealed no significant difference between the WT and YHR+ animals. Similar findings were found after quantification of apoptotic germ cell and Sertoli cells. Taken together, these data suggest that premature elevation in testosterone and persistently lower levels of circulating follicle stimulating hormone (FSH) are affecting Sertoli cell function, which is causing a reduced germ cell to Sertoli cell ratio in the YHR+ mice. These data suggest that the decrease in testis weight and seminiferous tubule diameter in YHR+ mine is due to a decrease in germ cell rather than Sertoli cell number.

The aim of this study was to examine the mechanisms of transcription and replication of the paramyxovirus, simian virus type 5 (SV5). This was initially attempted using reverse genetics techniques and subsequently examining specific viral protein: protein interactions within the replication complex. A cDNA clone encoding a synthetic negative-sense RNA genome analogue was constructed. Reverse genetics techniques were used to attempt to characterise conditions which supported the transcription and replication of this genome analogue, with or without the use of wild-type helper virus but were unsuccessful. During the course of these studies, a number of mammalian cell lines inducibly expressing SV5 proteins were isolated. These cell lines were subsequently used to examine viral protein: protein interactions within the replication complex. When expressed alone, both P and V proteins exhibited diffuse cytoplasmic fluorescence and V was also found in the nucleus. However, when NP was expressed alone, it was seen as punctate and granular cytoplasmic fluorescence. The distribution patterns of the proteins changed when expressed in combination. Large cytoplasmic aggregates similar to those at late times in an SV5 infection were seen in cells which co-expressed NP and P. When NP was co-expressed with V, however, NP was partially redistributed to give diffuse cytoplasmic and nuclear fluorescence. This showed that both P and V proteins could interact with NP and suggested that V may play a role in keeping NP soluble prior to an ordered encapsidation process. Extracts from these cell lines were then used in a novel protein: protein capture assay and demonstrated that NP could interact with both P and V proteins. NP expressed by the cell line was shown to contained both soluble and polymeric forms of NP. P was shown to bind both forms of NP, while V could only bind soluble NP. Since P and V proteins are amino co-terminal, the site of interaction between P and polymeric NP was predicted to be in the P unique C-terminus. This was strengthened when a P-specific C- terminal mAb was found to block the binding of P with polymeric NP. Deletion mutant analysis in the C-terminus of the P protein showed that the mAb binding site was at the extreme C-terminus of the protein suggesting this is the point of interaction between P and polymeric NP. Possible roles for these protein: protein interactions and implications for the paramyxovirus replication complex are discussed.

Large individuals of the sea anemone <i>Actinia fragacea</i> were collected at approximately monthly intervals over a two year period. Their gonads were examined by light and electron microscopy, in order to follow the gametogenic process. The sexes are separate, and both show an annual cycle of activity. Oocytes arise in the gonad epithelia, but soon migrate into the mesogloea. During vitellogenesis, the oocytes accumulate compound yolk granuleso fibrillar and cortical granules, lipid droplets and glycogen. The surface of large oocytes bears tufts of large microvilli or cytospines. The oocytes reach some 150 pm in size. A group of specialized gonad epithelial cells projects through the mesogloea and contacts the oocyte surface, forming the trophonema, which is involved with nutrient transfer. The gonad epithelium can take up nutrients from the external medium, and the trophonema is particularly active in the uptake and incorporation of some small molecules. Not all fully grown oocytes are always spawned; some break down in an orderly fashion and are resorbed. Oogenesis was also examined, in less detallo in four other species of anemone. Spermatogenesis takes place in spherical testicular cysts, which are also associated with trophonemata. Spawning occurred in the laboratory on three occasions. Spawned eggs do not possess a vitelline coat, and do not undergo a cortical reaction. Gastrulae may take up numerous supernumary sperm.

The Reproductive Homeobox X-linked, Rhox, genes encode transcription factors that are expressed exclusively in the testis, epididymis, placenta, and ovary. While there are 33 Rhox genes in mice, only Rhox5 and Rhox8 are expressed in Sertoli cells, suggesting that they alone regulate the expression of somatic-cell gene products crucial for testicular metabolism and germ-cell development. Targeted deletion of Rhox5, the founding member of the Rhox gene cluster, in mice, results in decreased expression of the insulin-2 gene (Ins2) and other metabolic genes, male subfertility via reduced sperm number, increased germ-cell apoptosis and a reduced proportion of sperm with normal motility. Davis et al. developed a Rhox8 siRNA knockdown transgenic model to study possible functional similarities between Rhox5 and Rhox8 and reveal compensatory actions via the breeding of Rhox5/Rhox8 double knockout mice. They observed that loss of Rhox8 results in downregulation of the sex-determining region Y gene (Sox9). Further analysis of the role of Rhox5 in testicular metabolism regulation was completed by development of mutant constructs encoding combinations of Rhox5 functional domains and subsequent analysis via qRT-PCR, luciferase assay and immunohistochemistry in cell lines transfected with expression plasmids containing these mutants. Our results indicated direct interaction of RHOX5 with the Ins2 promoter. The homeodomain and amino-terminal domain construct being sufficient for promoter activation albeit at a lower level than the full-length RHOX5 construct. MacLean et al. conducted qRT-PCR analysis of cells transfected with plasmids encoding the other Rhox genes revealed that Rhox8 and Rhox11 were also capable of upregulating Ins2 expression at a lower level than Rhox5. Our analysis of metabolic gene expression in the Rhox8 knockdown model also revealed decreased expression of Ins2 as well as insulin receptor-1 (InsR1). Continued analysis of the Rhox8-KD model and Rhox5/Rhox8 double knockout mice in young and aged (~12 months) male mice revealed a subfertility phenotype characterized by reduced litter frequency and size, reduced total spermatozoa and reduced sperm forward motility. This was reflected by a decrease in RHOX8 and SOX9 protein expression maintained in the aged mice that was more severe in the double knockout animals. qRT-PCR analysis of altered gene expression in the Rhox8 single knockdown male mice revealed significant expression downregulation of fellow Rhox genes Rhox5 and Rhox10 and expression upregulation of the growth differentiation factor-9 (Gdf9) gene normally expressed in the somatic cells of the ovary. Based on the regulation of sex specific genes Sox9 and Gdf9 in the Rhox8 knockdown model as well as data published by Daggag et al. revealing that Rhox8 is the only Rhox member expressed in the somatic cells of the embryonic testis, we sought to elucidate the possible role of Rhox8 in sex determination and testicular differentiation/development. Due to the limitations of our current knockdown model whereby siRNA production is initiated postnatally due to its androgen-dependent promoter, we elected to develop a new Rhox8 knockdown model to characterize the embryonic functions of Rhox8. Rhox8 siRNA were cloned into a Cre-recombinase activated expression vector in which a highly active U6 promoter drives expression of the shRNA. The constitutively active form of this vector exhibited knockdown of Rhox8 in stable Rhox8 overexpressing cell lines developed for this purpose. This developed vector is being used to develop a new transgenic line. While waiting for the development of the new Rhox8 knockdown transgenic mice, we chose to characterize the mRNA and protein expression of Rhox8 in the embryonic testis. Using qRT-PCR and immunohistochemistry, expression of Rhox8 was confirmed and exhibited continually increased expression in the Sertoli cells over the course of embryonic day ~13.5-18.5 (E13.5-E18.5). To examine possible functions of Rhox8 in the embryonic testis prior to receipt of the new knockdown model, we adapted a protocol to transfect the new constitutively active Rhox8 knockdown into embryonic gonads and cultured them up to 72 hours after electroporation. This protocol yielded successful expression of GFP from a GFP-expression plasmid and this was maintained for up to 72 hours in E13.5 gonads. qRT-PCR analysis of gonads transfected with the Rhox8 knockdown expression plasmid revealed significant knockdown of Rhox8 and Sox9 mRNA expression at the 48-hour and 72-hour time points.

The aim of the thesis was to analyse the reproductive efficiency of breeding geese in Fisheries Nové Hrady s.r.o. for the established period. The biggest difference in terms of fertilized eggs (P < 0.01) was observed between the objects of the Lomský předkmeny and Karolínský. A statistically significant difference of 13.42 % (P < 0.05) was among the objects of the Lomský předkmeny and Byňovský. The difference of fertilized eggs between the years 2016 and 2017 was of 25.64%, statistically highly significant (P < 0.01). The highest hatchability of setting eggs was detected in the object Karolínský. Was about of 15.01 % (P < 0.01) higher than in the object Lomský předkmeny, respectively. Byňovský (36.52%). In 2017 there was a large increase in hatchability of setting eggs of 24.95 % (P < 0.01), in comparison with the year 2016. The results of the hatchability of fertile eggs were similar. The biggest difference was recorded between the objects Byňovský and Karolínský. Difference 5.06% was not statistically significant. Differences in hatchability of fertile eggs have been between the object Lomský předkmeny and Karolínský, respectively Byňovský around 2.5 %. From the point of view, hatchability of fertile eggs was between the year 2016 and 2017, a statistically significant difference (7.48%, P < 0.01). The highest fertility of eggs were found in March, in April dropped by 15.82%, in May dropped by 11.60% and in June the decrease was only about 1.19%. A statistically significant effect (P < 0.05) were confirmed between fertility in March and May or June. Towards the end of the laying period by reducing the biological value of eggs occurred and to reduce the hatchability of setting eggs. Hatchability of setting eggs decreased in April by 13.75%, in May of 9.52% and in June only 0.78%. Differences in fertility between March and May or June was statistically significant (P < 0.05). For two-years-old geese, compared with one-years-old geese, fertility eggs fell by 29.1%, for three-years-old geese increased by 34.24% and four-years-old geese have declined by 18.75%. The difference between the two-years-old geese and one-years-old geese, respectively, 3-years-old geese were statistically significant (P < 0.05).

abstract: Pregnancy and childbirth are both natural occurring events, but still little is known about the signaling mechanisms that induce contractions. Throughout the world, premature labor occurs in 12% of all pregnancies with 36% of infant deaths resulting from preterm related causes. Even though the cause of preterm labor can vary, understanding alternative signaling pathways, which affect muscle contraction, could provide additional treatment options in stopping premature labor. The uterus is composed of smooth muscle, which is innervated, with a plexus of nerves that cover the muscle fibers. Smooth muscle can be stimulated or modulated by many sources such as neurotransmitters [i.e. dopamine], hormones [i.e. estrogen], peptides [i.e. oxytocin] and amines. This study focuses on the biogenic monoamine tyramine, which is produced in the tyrosine catecholamine biosynthesis pathway. Tyramine is known to be associated with peripheral vasoconstriction, increased cardiac output, increased respiration, elevated blood glucose and the release of norepinephrine. This research has found tyramine, and its specific receptor TAAR1, to be localized within mouse uterus and that this monoamine can induce uterine contractions at levels similar to oxytocin. / Dissertation/Thesis / Masters Thesis Biology 2017