Parthenogenetic activation of bovine oocytes using bovine and murine phospholipase C zeta

Background - During natural fertilization, sperm fusion with the oocyte induces long lasting intracellular calcium oscillations which in turn are responsible for oocyte activation. PLCZ1 has been identified as the factor that the sperm delivers into the egg to induce such a response. We tested the hypothesis that PLCZ1 cRNA injection can be used to activate bovine oocytes. Results - Mouse and bovine PLCZ1 cRNAs were injected into matured bovine oocytes at different concentrations. Within the concentrations tested, mouse PLCZ1 injection activated bovine oocytes at a maximum rate when the pipette concentration of cRNA ranged from 0.25 to 1 μg/μL, while bovine PLCZ1 was optimal at 0.1 μg/μL. At their most effective concentrations, PLCZ1 induced parthenogenetic development at rates similar to those observed using other activation stimuli such as Ionomycin/CHX and Ionomycin/DMAP. Injection of mouse and bovine PLCZ1 cRNA induced dose-dependent sperm-like calcium oscillations whose frequency increased over time. Injection of bovine and mouse PLCZ1 cRNA also induced IP3R-1 degradation, although bovine PLCZ1 cRNA evoked greater receptor degradation than its mouse counterpart. Conclusion - Injection of PLCZ1 cRNA efficiently activated bovine oocytes by inducing a sperm-like calcium oscillatory pattern. Importantly, the high rate of aneuploidy encountered in parthenogenetic embryos activated by certain chemical means was not observed in PLCZ1 activated embryos

Infrared seeded parametric four-wave mixing for sensitive detection of molecules

We have developed a sensitive resonant four-wave mixing technique based on two-photon parametric four-wave mixing with the addition of a phase matched ''seeder'' field. Generation of the seeder field via the same four-wave mixing process in a high pressure cell enables automatic phase matching to be achieved in a low pressure sample cell. This arrangement facilitates sensitive detection of complex molecular spectra by simply tuning the pump laser. We demonstrate the technique with the detection of nitric oxide down to concentrations more than 4 orders of magnitude below the capability of parametric four-wave mixing alone, with an estimated detection threshold of 10(12) molecules/cm(3)

Sensitive detection of sodium in a flame using parametric four-wave mixing and seeded parametric four-wave mixing

Two-photon resonant parametric four-wave mixing and a newly developed variant called seeded parametric four-wave mixing are used to detect trace quantities of sodium in a flame. Both techniques are simple, requiring only a single laser to generate a signal beam at a different wavelength which propagates collinearly with the pump beam, allowing efficient signal recovery. A comparison of the two techniques reveals that seeded parametric four-wave mixing is more than two orders of magnitude more sensitive than parametric four-wave mixing, with an estimated detection sensitivity of 5 x 10(9) atoms/cm(3). Seeded parametric four-wave mixing is achieved by cascading two parametric four-wave mixing media such that one of the parametric fields generated in the first high-density medium is then used to seed the same four-wave mixing process in a second medium in order to increase the four-wave mixing gain. The behavior of this seeded parametric four-wave mixing is described using semiclassical perturbation theory. A simplified small-signal theory is found to model most of the data satisfactorily. However, an anomalous saturationlike behavior is observed in the large signal regime. The full perturbation treatment, which includes the competition between two different four-wave mixing processes coupled via the signal field, accounts for this apparently anomalous behavior

Disciplina radnje, evokacija s fusnotama uloga vlaka, u školovanju

Tion. b) Pipette advanced into the oocyte; cytoplasm is aspirated to break the plasma membrane. c) Aspirated cytoplasm and Texas Red dextran are injected into the oocyte. d) Schematic representation of the microscope reticulum used as guide to control the injected volume. The oocyte is represented in yellow and the pipette in blue. The red lines indicate the volume introduced into the oocyte which, calculated measuring the pipette internal diameters at both ends, is 5.9 pL. e) An oil drop of the same size as the injected volume is shown next to an oocyte. f) Oocytes injected using Texas Red dextran. g) From left to right, oocyte injected 2X and 1X the normal volume of Texas Red dextran. h) Fluorescent intensity profile of the line shown in f. i) Fluorescent intensity profile of the line shown in g. j) Developmental rates of injected and uninjected bovine oocytes after activation using ionomycin/DMAP.<p><b>Copyright information:</b></p><p>Taken from "Parthenogenetic activation of bovine oocytes using bovine and murine phospholipase C zeta"</p><p>http://www.biomedcentral.com/1471-213X/8/16</p><p>BMC Developmental Biology 2008;8():16-16.</p><p>Published online 19 Feb 2008</p><p>PMCID:PMC2266721.</p><p></p

Fertilization induces a transient exposure of phosphatidylserine in mouse eggs

Phosphatidylserine (PS) is normally localized to the inner leaflet of the plasma membrane and the requirement of PS translocation to the outer leaflet in cellular processes other than apoptosis has been demonstrated recently. In this work we investigated the occurrence of PS mobilization in mouse eggs, which express flippase Atp8a1 and scramblases Plscr1 and 3, as determined by RT-PCR; these enzyme are responsible for PS distribution in cell membranes. We find a dramatic increase in binding of flouresceinated-Annexin-V, which specifically binds to PS, following fertilization or parthenogenetic activation induced by SrCl2 treatment. This increase was not observed when eggs were first treated with BAPTA-AM, indicating that an increase in intracellular Ca2+ concentration was required for PS exposure. Fluorescence was observed over the entire egg surface with the exception of the regions overlying the meiotic spindle and sperm entry site. PS exposure was also observed in activated eggs obtained from CaMKIIγ null females, which are unable to exit metaphase II arrest despite displaying Ca2+ spikes. In contrast, PS exposure was not observed in TPEN-activated eggs, which exit metaphase II arrest in the absence of Ca2+ release. PS exposure was also observed when eggs were activated with ethanol but not with a Ca2+ ionophore, suggesting that the Ca2+ source and concentration are relevant for PS exposure. Last, treatment with cytochalasin D, which disrupts microfilaments, or jasplakinolide, which stabilizes microfilaments, prior to egg activation showed that PS externalization is an actin-dependent process. Thus, the Ca2+ rise during egg activation results in a transient exposure of PS in fertilized eggs that is not associated with apoptosis.Fil: Curia, Claudio Augusto. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); ArgentinaFil: Ernesto, Juan Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); ArgentinaFil: Stein, Paula. University of Pennsylvania; Estados UnidosFil: Busso, Dolores. Pontificia Universidad Católica de Chile; ChileFil: Schultz, Richard. University of Pennsylvania; Estados UnidosFil: Cuasnicu, Patricia Sara. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); ArgentinaFil: Cohen, Debora Juana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentin

Short Staple Variety Demonstration Summary, Maricopa Agricultural Center, 1989-1991

Short staple variety trials have been conducted for three years on the demonstration farm, at the Maricopa Agricultural Center. Twenty varieties, received from various seed companies, were entered into the test in 1989, 19 varieties in 1990, and 10 varieties in 1991. Yields ranged from 3353 to 4679 lbs. seedcotton/acre in 1989, 829 to 1428 lbs. lint/acre in 1990 and 1099 to 1246 lbs. lint/acre in 1991. Results form the statistical analysis showed significant differences among varieties for all three years. Differences among varieties in 1989 showed no clear trends with respect to maturity types. In 1990, those varieties that were medium to early season maturity types yielded higher than full season maturity types. The results in 1991 showed medium maturity types yielding in the highest ranking of the test without exception, while the performance of the full season maturity types was mixed. An additional test with narrow row spacing (30 inches) was included in 1991 with 5 varieties. Yields ranged from 947 to 1117 lbs. lint/acre. No significant differences were detected among varieties

Short Staple Variety Demonstration, Maricopa Agricultural Center, 1990

A short staple variety trial was conducted on the demonstration farm, at the Maricopa Agricultural Center in 1989. Nineteen varieties, received from various seed companies, were entered into the test. Yields ranged from 829 to 1428 lbs. lint acre⁻¹ for STY 110 and STV 1324 respectively. Results from the statistical analysis showed significant differences among varieties. Those varieties that were medium to short - season maturity types yielded higher than long, full season maturity types. This difference in yield was most probably due to the hot and humid weather conditions experienced in the 1990 growing season and the varietal differences associated with heat tolerance

Cottonseed Treatment Evaluations in Arizona, 1990

Field experiments were conducted at four locations in Arizona (Yuma, Maricopa, Marana, Safford) to evaluate 16 cottonseed treatments on cotton that included 12 on Upland (Q. hirsutum L.) and 4 on Pima (Gossvpium barbadense L.). Stand counts were taken to evaluate the effectiveness of each treatment. Statistical analysis showed no significant differences among the treatments used for the Upland cottonseed. Significant differences were found among the treatments used for the Pima cotton seed at the Marana and Safford locations only