【摘要】：本研究的主要目的是探索低溫處理對(duì)豬精子蛋白泛素化水平及體外受精的影響。本試驗(yàn)主要采用CASA、SDS-PAGE、Western blot等試驗(yàn)方法進(jìn)行分析,得出如下結(jié)果：1.常溫、冷休克、低溫處理后附睪精子活率、頂體完整性還是質(zhì)膜完整性均高于精子試驗(yàn)組。2.常溫、冷休克、低溫處理后附睪精子的平均曲線運(yùn)動(dòng)速度(VCL)、平均直線運(yùn)動(dòng)速度(VSL)、平均路徑運(yùn)動(dòng)速度(VAP)均高于精子試驗(yàn)組。3.常溫、冷休克、低溫處理后鮮精和附睪精子的蛋白進(jìn)行SDS-PAGE,實(shí)驗(yàn)結(jié)果顯示,冷休克和低溫后的豬精子蛋白含量或種類發(fā)生了變化,而鮮精與附睪精子樣本基本沒有差異。4.冷休克處理的豬精子樣本進(jìn)行Western blot分析。實(shí)驗(yàn)結(jié)果表明：得知約在30kDa、46kDa和62kDa處存在泛素分子：對(duì)46kDa處進(jìn)行灰度分析,各處理組之間條帶的灰度值比都差異明顯；62kDa處只在鮮精和附睪精子中發(fā)現(xiàn)泛素分子；而冷休克處理的泛素化程度相對(duì)其它兩處理組降低很多。5.低溫處理的豬精子樣本進(jìn)行Western blot分析。實(shí)驗(yàn)結(jié)果表明：得知約在46kDa和62kDa處存在泛素分子,對(duì)46kDa處的泛素化程度進(jìn)行灰度分析,精子-80℃處理組與其它三個(gè)處理組之間的灰度比值都小很多；附睪精子-196℃處理組與其它三個(gè)處理組之間的灰度比值都大很多。6.冷休克處理的豬精子與成熟卵母細(xì)胞進(jìn)行體外受精,受精2d后觀察卵裂情況,并統(tǒng)計(jì)卵裂率。實(shí)驗(yàn)結(jié)果表明：附睪鮮精卵裂率高于鮮精；冷休克附睪精子卵裂率高于冷休克精子。7.低溫處理的豬精子與成熟卵母細(xì)胞進(jìn)行體外受精,受精2d后觀察卵裂情況,并統(tǒng)計(jì)卵裂率。實(shí)驗(yàn)結(jié)果表明：附睪精子-80℃卵裂率高于精子-80℃；附睪精子-196℃的卵裂率高于精子-196℃。
[Abstract]:The aim of this study was to explore the effects of low temperature treatment on the level of ubiquitin and in vitro fertilization of porcine sperm. The results of this experiment are as follows: 1. The results are as follows: 1. CASA,SDS-PAGE,Western blot and other test methods are used for analysis. The sperm motility, acrosome integrity or plasma membrane integrity of epididymal spermatozoa after normal temperature, cold shock and hypothermic treatment were higher than those of sperm treatment group. 2. The mean curvilinear velocity (VCL),) of epididymal sperm after normal temperature, cold shock and hypothermic treatment was higher than that of sperm treated with low temperature (VSL), mean linear velocity (VSL), mean path velocity (VAP). The results of SDS-PAGE, test of fresh sperm and epididymal spermatozoa after cold shock and cold shock showed that the protein content or species of pig spermatozoa changed after cold shock and hypothermic treatment. There was no difference between fresh sperm and epididymal sperm samples. 4. Cold shock-treated pig sperm samples were analyzed by Western blot. The results showed that there were ubiquitin molecules at about 30kDas, 46kDa and 62kDa. The gray-scale ratio of the bands at 46kDa was significantly different among the treatment groups, and only ubiquitin molecules were found in fresh sperm and epididymal sperm in 62kDa, while ubiquitin molecules were only found in fresh sperm and epididymal spermatozoa in 62kDa. The degree of ubiquitin of cold shock treatment was much lower than that of the other two groups. 5. 5%. Low temperature-treated pig sperm samples were analyzed by Western blot. The results showed that there were ubiquitin molecules in 46kDa and 62kDa, and the degree of ubiquitin at 46kDa was analyzed by gray scale analysis. The gray scale ratio between sperm-80 鈩，

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