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1.
试验旨在探讨单精子胞浆内注射(intracytoplasmic sperm injection,ICSI)法生产猪体外受精卵和应用猪ICSI受精卵进行胞质注射生产转基因胚胎的可行性。首先对比了猪体外受精(in vitro fertilization,IVF)受精卵与ICSI受精卵的胚胎发育效率;然后观察了猪ICSI受精卵的双原核形成时间及效率,对精子注射到胞质后6~18h分6个时间段进行地衣红染色,对比精子进入卵胞质后的状态及原核形成;最后对猪IVF受精卵受精后8~10h及ICSI受精卵受精后12~14h进行EGFP-N1质粒(20ng/μL)胞质注射,观察胚胎发育效率及转基因效率。结果表明,ICSI受精卵的胚胎发育率(卵裂率89.4%和67.9%、囊胚率36.5%和16.1%)显著优于IVF组(P0.05),适合用于猪的体外受精卵试验;猪ICSI受精卵双原核在精子注射到卵胞质后12~14h形成,双原核形成率为54.90%,显著高于其余5个试验组(P0.05);ICSI受精卵胞质注射组胚胎卵裂率(86.2%和66.3%)、囊胚率(30.0%和13.6%)及转基因效率(18.5%和0)均显著高于IVF受精卵胞质注射试验组(P0.05)。本试验结果为采用ICSI受精卵进行胞质注射生产转基因猪的研究奠定了基础。 相似文献
2.
本试验探讨了不同辅助激活方法(Calciumionophore A23187激活、Calciumionophore A23187+6-DMAP联合激活和电激活)、不同精子预处理方法(液氮冻融处理和0.1%Triton X-100处理)和在添加半胱氨酸的胚胎培养液中培养不同时间(0 h、4 h、12 h和168 h)对猪卵母细胞内单精子注射(ICSI)胚胎体外发育的影响。结果显示:与无辅助激活相比,A23187+6-DMAP联合激活和电激活均能显著提高ICSI卵母细胞的激活率、卵裂率和囊胚率(P0.05),A23187+6-DMAP联合激活能显著提高ICSI卵母细胞的受精率(P0.05)。液氮冻融精子组ICSI卵母细胞的雄原核形成率显著高于活精子组(P0.05)。在添加半胱氨酸的胚胎培养液中培养4 h的ICSI卵母细胞受精率、雄原核形成率和囊胚率显著高于0 h组(P0.05)。以上结果表明,猪卵母细胞在ICSI后需要辅助激活来启动胚胎顺利发育,A23187+6-DMAP激活效果较好。液氮冻融精子可以促进ICSI后雄原核的形成。半胱氨酸处理4 h对猪ICSI卵母细胞受精和发育均有促进作用。 相似文献
3.
为探讨猪精子、水牛精子及食蟹猴精子分别注入到猪卵母细胞后原核形成及早期胚胎发育情况,利用屠宰场收集的猪卵母细胞,经体外成熟44～48h后,进行胞质内显微受精(ICSI)操作。试验1:体外培养18～19h,用Ho-echst33342荧光染色,检查原核形成情况。试验2:进行体外培养,ICSI后2d检查分裂率,7d记录囊胚率。试验1结果显示:在原核形成率上,猪同种显微受精,原核形成率(50.10%)显著高于注入水牛精子(37.06%)及食蟹猴精子(37.48%),且差异显著(P0.05),注入水牛精子和食蟹猴精子间差异不显著。试验2结果显示:猪同种显微受精所得的分裂率(86.58%)和囊胚率(29.93%)与水牛精子注入(70.98%,18.48%)、食蟹猴精子注入(78.69%,16.92%)差异显著(P0.05)。结果表明,水牛精子、食蟹猴精子分别注入到猪卵母细胞后,观察到雌雄原核和精子解聚;水牛精子注入猪卵母细胞与食蟹猴精子注入猪卵母细胞,经体外培养发育到囊胚。 相似文献
4.
试验旨在探讨单精子胞浆内注射（intracytoplasmic sperm injection,ICSI）法生产猪体外受精卵和应用猪ICSI受精卵进行胞质注射生产转基因胚胎的可行性。首先对比了猪体外受精（in vitro fertilization,IVF）受精卵与ICSI受精卵的胚胎发育效率;然后观察了猪ICSI受精卵的双原核形成时间及效率,对精子注射到胞质后6~18 h分6个时间段进行地衣红染色,对比精子进入卵胞质后的状态及原核形成;最后对猪IVF受精卵受精后8~10 h及ICSI受精卵受精后12~14 h进行EGFP-N1质粒（20 ng/μL）胞质注射,观察胚胎发育效率及转基因效率。结果表明,ICSI受精卵的胚胎发育率（卵裂率89.4%和67.9%、囊胚率36.5%和16.1%）显著优于IVF组（P<0.05）,适合用于猪的体外受精卵试验;猪ICSI受精卵双原核在精子注射到卵胞质后12~14 h形成,双原核形成率为54.90%,显著高于其余5个试验组（P<0.05）;ICSI受精卵胞质注射组胚胎卵裂率（86.2%和66.3%）、囊胚率（30.0%和13.6%）及转基因效率（18.5%和0）均显著高于IVF受精卵胞质注射试验组（P<0.05）。本试验结果为采用ICSI受精卵进行胞质注射生产转基因猪的研究奠定了基础。 相似文献
5.
试验采用不同浓度的DTT或GSH预处理猪精子,统计ICSI后精核解聚率、雄原核形成率及胚胎发育情况.结果表明,①精核解聚率随DTT浓度的增加而提高,6 mmol/L DTT处理组的2PN 2Pb形成率(P＜0.01)和卵裂率(P＜0.05)均高于其余各组.随着DTT浓度的增加,囊胚总细胞数呈现下降趋势.表明6 mmol/LDTT预处理精子的ICSI效果较好,但DTT在促进精核解聚的同时所产生的副作用在一定程度上影响了ICSI胚胎的后期发育;②随着GSH浓度的升高,注射卵母细胞的2PN 2Pb形成率、MPN形成率和精核解聚率均呈上升趋势.0.5 mmol/L GSH处理组的卵裂率高于其余处理组(P＜0.05)和对照组(P＜0.01),囊胚发育率显著高于对照组(P＜0.05).说明0.50 mmol/L GSH处理组的ICSI效果较好;③0.5 mmol/L GSH处理组的2PN 2Pb形成率、卵裂率和囊胚总细胞数极显著高于6 mmol/LDTT处理组(P＜0.01),两者MPN形成率差异显著(P＜0.05).表明0.5 mmol/L GSH预处理精子的ICSI效果要好于6 mmol/L DTT. 相似文献
6.
试验探讨了水牛和黄牛同种及种间显微授精的可行性。体外成熟22~24 h的水牛和黄牛卵母细胞分别注入冷冻/解冻的尼里－拉菲水牛和利木赞黄牛精子,进行同种或种间的显微授精操作,构建的ICSI胚胎一部分培养到16~18 h固定染色检查原核形成情况,另外一部分胚胎培养2~9 d分别记录胚胎的分裂情况及囊胚发育情况。结果发现,水牛同种(水牛+水牛)和水牛异种(水牛+黄牛)显微授精胚胎的双原核率(47.73%和36.67%)、分裂率(68.00%和72.64%)和囊胚发育率(16.44%和22.39%)均无显著差异(P＞0.05);黄牛同种(黄牛+黄牛)和黄牛异种(黄牛+水牛)显微授精胚胎的双原核率(60.00%和45.28%)、分裂率(73.33%和71.31%)和囊胚发育率(28.57%和33.70%)亦无显著差异(P＞0.05)。以上结果表明:①水牛精子注射到黄牛卵子和黄牛精子注射到水牛卵子的种间授精胚胎均可以体外发育到囊胚阶段;②黄牛卵子无论同种和种间显微授精的效果均优于水牛卵子。 相似文献
7.
本研究旨在探讨不同的联合激活处理、精子预处理方法和单精子注射方式对奶牛分离精子ICSI效率的影响。借助显微操作仪将经不同预处理的奶牛分离X精子直接注入体外成熟22～24h的牛卵母细胞胞质内,注射过程采用回吸胞质和不回吸胞质2种处理,最后分别用3种激活方案对注射卵进行激活。结果表明:用CR1aa+Ionomycin+6-DMAP对ICSI注射卵进行激活后,卵裂率和囊胚率都高于A23187+CHX和7%ET+CHX两种联合激活组(63.74%vs61.09%、53.75%;28.54%vs17.68%、22.00%,P0.05);采用percoll密度梯度离心法处理精子,卵裂率(77.10%vs62.19%,P0.05)与囊胚率(26.95%vs22.82%,P0.05)均高于上游法处理组;ICSI时回吸胞质获得的注射卵,其卵裂率和囊胚率均显著高于不回吸胞质处理组(63.10%,25.35%vs41.56%,19.40%,P0.05)。结果表明,用percoll密度梯度离心法处理精子、注射时回吸胞质、用CR1aa+Iono-mycin+6-DMAP激活ICSI注射卵,可显著提高奶牛分离精子ICSI的效率。 相似文献
8.
系统探讨了几种激活方法对水牛卵母细胞孤雌发育的影响。体外成熟 (IVM) 2 6 h的水牛卵母细胞经 7%乙醇(EH)、钙离子载体 (A2 3187)或离子霉素 (Ion)激活处理 5 min后 ,在 2 m mol/ L 6 -甲二氨基嘌呤 (6 - DMAP)中培养 3～4 h,然后再继续培养 6～ 11d,观察其胚胎发育情况。结果发现 ,5μmol/ L Ion激活处理的水牛卵母细胞分裂率显著高于 EH处理的卵母细胞 (6 3.0 % vs5 4 .2 % ,P<0 .0 5 ) ,其原核形成率也显著高于 EH处理的卵母细胞。激活处理前 ,无Ca2 培养液孵育时间的长短 ,对水牛卵母细胞孤雌发育无显著影响 (P>0 .0 5 )。如用 A2 3187激活处理水牛卵母细胞 ,其激活分裂率则随着浓度的升高而提高。从而表明 ,5 μmol/ L Ion可有效激活水牛卵母细胞 相似文献
9.
实验研究了不同成熟培养时间的牛卵母细胞玻璃化冷冻及胞质内单精子注射(ICSI)后的受精效果。结果表明:成熟后的新鲜牛卵母细胞按照ICSI注射方法穿刺而不注射精子组与未经穿刺的对照组相比,孤雌激活后的卵裂率、囊胚发育率及囊胚细胞数无显著差异(P>0.05);成熟培养16h(MⅠ)和23h(MⅡ)卵母细胞冷冻解冻后形态正常率均显著低于新鲜对照组(76.66%、87.33%vs100.0%)(P<0.05),冷冻解冻后二者分别成熟培养至24h,ICSI后胚胎的囊胚发育率(5.29%、14.41%)显著低于新鲜对照组(24.40%)(P<0.05);成熟培养23h与成熟培养16h的卵母细胞冷冻解冻后形态正常率及ICSI后囊胚发育率(14.41%vs5.29%)均有显著性差异(P<0.05)。实验证明,ICSI操作不会影响卵母细胞发育潜力;玻璃化冷冻影响卵母细胞解冻后形态正常率以及ICSI后胚胎的发育能力;成熟培养23h比16h的卵母细胞冷冻保存后经ICSI的胚胎发育潜力高。 相似文献
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11.
The present study examines the contribution of the nucleus to meiotic competence in mouse oocytes that were reconstructed using nuclear transfer. Three types of reconstructed oocytes were produced: MP‐GV, by transplanting the male pronucleus (MP) into germinal vesicle (GV) stage oocytes; 3T3‐GV, by transplanting the nucleus of a National Institute of Health (NIH) 3T3 cell into a GV stage oocyte; and 3T3‐MII, by transplanting the nucleus of an NIH 3T3 cell into a metaphase II (MII) stage oocyte. The fusion rates differed, but not significantly, in the MP‐GV, 3T3‐GV, and 3T3‐MII groups (77, 63, 56%, respectively). Then, meiotic competence was compared in MP‐GV, 3T3‐GV and non‐manipulated GV stage oocytes as a control. Nuclear envelope breakdown occurred in all the reconstructed oocytes, as well as the control ones. The percentage of first polar body extrusion differed between the MP‐GV (100%), 3T3‐GV (72%), and control (67%) groups. DNA staining with Hoechst 33342 revealed that in the MP‐GV‐group oocytes that had reached MII stage, the chromosomes were condensed and aligned in a regular array similar to the normal metaphase plate. By contrast, in 3T3‐GV group oocytes, the condensed chromosomes were irregularly scattered in the cytoplasm. These results suggest that the donor nucleus affects meiotic competence in reconstructed oocytes. 相似文献
12.
Hiep Thi Nguyen Thanh Quang Dang-Nguyen Tamas Somfai Nguyen Thi Men Barbara Beck-Woerner Nguyen Viet Linh Bui Xuan Nguyen Junko Noguchi Hiroyuki Kaneko Kazuhiro Kikuchi 《Animal Science Journal》2021,92(1):e13650
Male pronucleus (MPN) formation is a very important physiological event during fertilization, which affects in vitro production of transferrable embryos. The aim of this study was to find out the correlation between the number of penetrated sperm and the occurrence of failure of MPN formation in porcine oocytes. In vitro matured porcine oocytes were fertilized in vitro with frozen epididymal sperm. Two different frozen sperm lots were tested in this study, which were different in terms of polyspermy rates. The numbers and the status of penetrated sperm in oocytes were evaluated 10 h after insemination. Under high polyspermy condition, the polyspermy rate was 83.5% with an average mean of 3.5 sperms per penetrated oocyte, whereas the percentage of polyspermy was 65.5% with an average mean of 2.4 sperms per penetrated oocyte under moderate polyspermic condition. Correlation analysis revealed a negative correlation between the number of penetrated sperm and their MPN formation percentage both in the sperm lot of high polyspermy (R = −0.560, p < 0.05) and in the sperm lot of moderate polyspermy (R = −0.405, p < 0.05) which suggests that penetration of excessive spermatozoa disables the oocyte cytoplasm to promote MPN formation. 相似文献
13.
This study was conducted to investigate the pattern of DNA methylation in pronuclearstage mouse embryos derived from vitrified-warmed oocytes.Mouse oocytes at metaphase Ⅱ (MⅡ) stage of meiosis were all... 相似文献
14.
试验选用96头平均体重14.82 kg左右的杜×长×大断奶仔猪,随机分成4组,每组3栏,每栏8头(公母各半)。对照组饲喂基础日粮,试验1、2、3组分别添加1%80目白术、0.2%白术多糖和1%微米白术。试验期30 d。结果表明:在生长性能方面,与对照组相比,1%微米白术添加组可显著提高日增重(P0.05)、降低饲料增重比和腹泻率,而且效果优于1%80目白术组和0.2%白术多糖组,在肠道形态和肠道微生态区系方面,与对照组相比,日粮添加1%80目白术、0.2%白术多糖、1%微米白术均可不同程度的提高十二指肠和空肠的绒毛高度,加深十二指肠和空肠的隐窝深度,并且增加肠道微生态区系的多样性,其中以1%微米白术添加组的效果最佳。 相似文献
15.
Sissay, M.M., Uggla, A. and Waller, P.J., XXXX. Prevalence and seasonal incidence of nematode parasites and fluke infections
of sheep and goats in eastern Ethiopia. Tropical Animal Health and Production, XXXX. A 2-year abattoir survey was carried out to determine the prevalence, abundance and seasonal incidence of gastro-intestinal
(GI) nematodes and trematodes (flukes) of sheep and goats in the semi-arid zone of eastern Ethiopia. During May 2003 to April
2005, viscera including liver, lungs and GI tracts were collected from 655 sheep and 632 goats slaughtered at 4 abattoirs
located in the towns of Haramaya, Harar, Dire Dawa and Jijiga in eastern Ethiopia. All animals were raised in the farming
areas located within the community boundaries for each town. Collected materials were transported within 24 h to the parasitology
laboratory of Haramaya University for immediate processing. Thirteen species belonging to 9 genera of GI nematodes (Haemonchus contortus, Trichostrongylus axei, T. colubriformis, T. vitrinus, Nematodirus filicollis, N. spathiger, Oesophagostomum
columbianum, O. venulosum, Strongyloides papillosus, Bunostomum trigonocephalum, Trichuris ovis, Cooperia curticei and Chabertia ovina), and 4 species belonging to 3 genera of trematodes (Fasciola hepatica, F. gigantica, Paramphistomum {Calicohoron} microbothrium and Dicrocoelium dendriticum) were recorded in both sheep and goats. All animals in this investigation were infected with multiple species to varying
degrees. The mean burdens of adult nematodes were generally moderate in both sheep and goats and showed patterns of seasonal
abundance that corresponded with the bi-modal annual rainfall pattern, with highest burdens around the middle of the rainy
season. In both sheep and goats there were significant differences in the mean worm burdens and abundance of the different
nematode species between the four geographic locations, with worm burdens in the Haramaya and Harar areas greater than those
observed in the Dire Dawa and Jijiga locations. Similar seasonal variations were also observed in the prevalence of flukes.
But there were no significant differences in the prevalence of each fluke species between the four locations. Overall, the
results showed that Haemonchus, Trichostrongylus, Nematodirus, Oesophagostomum, Fasciola and Paramphistomum species were the most abundant helminth parasites of sheep and goats in eastern Ethiopia. 相似文献
16.
Tropical Animal Health and Production - 相似文献
17.
Rassnick KM Gieger TL Williams LE Ruslander DM Northrup NC Kristal O Myers NC Moore AS 《Journal of veterinary internal medicine / American College of Veterinary Internal Medicine》2001,15(3):196-199
1-(2-Chloroethyl)3-cyclohexyl-1-nitrosourea (CCNU) is an alkylating agent in the nitrosourea subclass. A prospective evaluation of CCNU was done to determine the maximally tolerated dosage of CCNU in tumor-bearing cats. Response data were obtained when available. Twenty-five cats were treated with CCNU at a dosage of 50-60 mg/m3 body surface area. Complete hematologic data were available for 13 cats. Neutropenia was the acute dose-limiting toxicity. The median neutrophil count at the nadir was 1,000 cells/microL (mean, 2,433 cells/microL; range, 0-9,694 cells/microL). The time of neutrophil nadir was variable, occurring 7-28 days after treatment, and counts sometimes did not return to normal for up to 14 days after the nadir. Based on these findings, a 6-week dosing interval and weekly hematologic monitoring after the 1st treatment with CCNU are recommended. The nadir of the platelet count may occur 14-21 days after treatment. The median platelet count at the nadir was 43,500 cells/microL. No gastrointestinal, renal, or hepatic toxicities were observed after a single CCNU treatment, and additional studies to evaluate the potential for cumulative toxicity should be performed. Five cats with lymphoma and 1 cat with mast cell tumor had measurable responses to CCNU. Phase II studies to evaluate antitumor activity should be completed with a dosing regimen of 50-60 mg/m3 every 6 weeks. 相似文献
18.
H Hayashi T Yoshimura J Y Chen 《Comparative immunology, microbiology and infectious diseases》1985,8(2):73-87
Our particular attention in this article was given to natural mediators for macrophages isolated from the sites of tissue injury. A number of chemotactic factors, which may satisfy many criteria making them acceptable as inflammatory leucocyte chemotactic factors, has been separated. Among them, our laboratory has isolated three macrophage (monocyte) chemotactic factors (MCF-a, -b and -c). Their purification, characterization and functional specificity are discussed. 相似文献
19.
Furosemide continuous rate infusion in the horse: evaluation of enhanced efficacy and reduced side effects 总被引：1，自引：0，他引：1
Johansson AM Gardner SY Levine JF Papich MG LaFevers DH Fuquay LR Reagan VH Atkins CE 《Journal of veterinary internal medicine / American College of Veterinary Internal Medicine》2003,17(6):887-895
Continuous rate infusion (CRI) of furosemide in humans is considered superior to intermittent administration (IA). This study examined whether furosemide CRI, compared with IA, would increase diuretic efficacy with decreased fluid and electrolyte fluctuations and activation of the renin-angiotensin-aldosterone system (RAAS) in the horse. Five mares were used in a crossover-design study. During a 24-hour period, each horse received a total of 3 mg/kg furosemide by either CRI (0.12 mg/kg/h preceded by a loading dose of 0.12 mg/kg IV) or IA (1 mg/kg IV q8h). There was not a statistically significant difference in urine volume over 24 hours between methods; however, urine volume was significantly greater after CRI compared with IA during the first 8 hours ([median 25th percentile, 75th percentile]: 9.6 L [8.9, 14.4] for CRI versus 5.9 L [5.3, 6.0] for IA). CRI produced a more uniform urine flow, decreased fluctuations in plasma volume, and suppressed renal concentrating ability throughout the infusion period. Potassium, Ca, and Cl excretion was greater during CRI than IA (1,133 mmol [1.110, 1,229] versus 764 mmol [709, 904], 102.7 mmol [96.0, 117.2] versus 73.3 mmol [65.0, 73.5], and 1,776 mmol [1,657, 2.378] versus 1,596 mmol [1,457, 1,767], respectively). Elimination half-lives of furosemide were 1.35 and 0.47 hours for CRI and IA, respectively. The area under the excretion rate curve was 1,285.7 and 184.2 mL x mg/mL for CRI and IA, respectively. Furosemide CRI (0.12 mg/kg/h) for 8 hours, preceded by a loading dose (0.12 mg/kg), is recommended when profound diuresis is needed acutely in horses. 相似文献
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B. Cinar Kul N. Bilgen J.A. Lenstra O. Korkmaz Agaoglu B. Akyuz O. Ertugrul 《Zeitschrift für Tierzüchtung und Züchtungsbiologie》2015,132(6):449-453
Genetic variations in chromosome Y are enabling researchers to identify paternal lineages, which are informative for introgressions and migrations. In this study, the male‐specific region markers, sex‐determining region‐Y (SRY), amelogenin (AMELY) and zinc finger (ZFY) were analysed in seven Turkish native goat breeds, Angora, Kilis, Hair, Honaml?, Norduz, Gürcü and Abaza. A SNP in the ZFY gene defined a new haplotype Y2C. All domestic haplogroups originate from Capra aegagrus, while the finding of Y1A, Y1B, Y2A and Y2C in 32, 4, 126 and 2 Turkish domestic goats, respectively, appears to indicate a predomestic origin of the major haplotypes. The occurrence of four haplotypes in the Hair goat and, in contrast, a frequency of 96% of Y1A in the Kilis breed illustrate that Y‐chromosomal variants have a more breed‐dependent distribution than mitochondrial or autosomal DNA. This probably reflects male founder effects, but a role in adaptation cannot be excluded. 相似文献