Optimizing injection time of GFP plasmid into sheep zygote

Microinjection of exogenous DNA into the cytoplasm of matured oocytes or zygotes is a promising technique to generate transgenic animals. However, the data about the microinjection time and procedure in sheep are limited and have not treated in detail. To obtain more in-depth information, the Sarda sheep oocytes from abattoir-derived ovaries were subjected to IVM and IVF. Then, the GFP plasmid as a reporter gene was injected into the cytoplasm of MII oocytes (n: 95) and zygotes at different post-insemination intervals (6–8 hpi, n: 120; 8–10 hpi, n: 122; 10–12 hpi, n: 110 and 12–14 hpi, n: 96). There were no significant differences in the cleavage rates between the groups. However, blastocyst rate of injected zygotes at all-time intervals was significantly lower than injected MII oocytes and control group (p < 0.05). Interestingly, the proportion of GFP-positive embryos was higher at 8–10 hpi compared with other injected groups (4 % versus 0 %, p  < 0.01). Among these, the proportion of mosaic embryos was high and two of those embryos developed to the blastocyst stage. In conclusion, we settled on the cytoplasmic microinjection of GFP plasmid at 8–10 hpi as an optimized time point for the production of transgenic sheep and subsequent experiments.     

Gluten Structural Evolution During Pasta Processing of Refined and Whole Wheat Pasta from Hard White Winter Wheat: The Influence of Mixing,Drying, and Cooking

An attempt was made to evaluate gluten structural changes in refined and whole wheat pasta from hard white winter wheat to elucidate the impact of whole wheat components on the formation and structure of the gluten network in pasta. Attenuated total reflectance–FTIR spectroscopy was used to track gluten secondary structure through most of the major steps in pasta processing: raw material, mixing, drying, and cooking. Protein solubility, accessible thiols, and SDS‐PAGE data were also collected to provide additional information on the nature of protein interactions and network composition. Few secondary structural differences were observed between refined and whole wheat flours from hard white wheat. However, mixing induced a significant shift to β‐sheet structures in refined dough that was not equally matched by whole wheat dough. Drying under both high temperature, short time (HT) and low temperature, long time (LT) conditions resulted in a reversion to structural distributions similar to those for flour in both pastas. However, greater protein denaturation in HT samples was indicated by lower protein solubility also in the presence of denaturants and disulfide reducing agents. Cooking generated a substantial increase in β‐sheet structures for both pasta systems. This structure was greatest in refined and LT samples. Thiol accessibility data indicate the presence of a highly aggregated, compact gluten network in refined pasta, mostly driven by hydrophobic association. Conversely, the network in whole wheat pasta was more loosely associated and dependent on disulfide bonding, both of which fit well with the secondary structural data.     

Clinical presentation and management of suspected ribavirin toxicosis in a dog

A 5-month-old pit bull terrier was presented for evaluation of progressive lethargy, vomiting, diarrhea, and anorexia 45 hours after ingestion of 625 mg/kg body weight (BW) (9000 mg) of the antiviral medication, ribavirin. Abnormalities that were detected included dehydration, tachycardia, elevated liver enzymes, and prolonged prothrombin time. The dog was discharged after 5 days of aggressive supportive care consisting of intravenous fluids, antiemetics, gastroprotectants, hepatoprotectants, dextrose supplementation, and vitamin B/K1 supplementation.