牦牛睾丸附睾LDH同工酶表达模式的研究

利用比色法和聚丙烯酰胺凝胶电泳(PAGE)技术对6、12、18、24月龄半血野牦牛、横交半血野牦牛和家牦牛睾丸、附睾组织LDH同工酶活性及谱带表达进行测析.结果表明睾九、附睾LDH在相同月龄其表达模式不同,6月龄睾丸LDH有4条酶谱,活性百分率依次(LDH1＞LDH3＞LDH2＞LDH4),附睾只有3条酶谱(LDH1＞LDH2＞LDH3);到12、18月龄时,睾丸LDH有5条酶谱(LDH1＞LDH3＞LDH2＞LDH2＞LDH4＞LDH5),附睾4条酶漕(LDH1＞LDH2＞LDH3＞LDH4),睾丸和附睾LDH同工酶随月龄增长和睾丸组织发育,活性和酶谱表现出明显组织器官的特异性.3种类型牦牛LDH同工酶表达基本相似.24月龄时,睾丸、附睾LDH有6条谱带,LDH-X位于LDH4与LDH5之间,证明此时牦公牛睾丸中精子成熟.     

青海双峰驼血液LDH和SOD同工酶酶谱

采用聚丙烯烯酰胺凝胶电泳法对５８峰青海双峰驼的红细胞和血清乳酸脱氢酶（ＬＤＨ）以及红细胞超氧化物歧化酶（ＳＯＤ）同工酶酶谱进行了分析和研究，结果：（１）ＳＬＤＨ同工酶酶谱存在年龄差异，成年驼只有ＬＤＨ１同工酶，幼年驼酶谱由ＬＤＨ１，ＬＤＨ２，ＬＤＨ３和ＬＤＨ４组成；（２）ＲＢＣ－ＬＤＨ同工酶由ＬＤＨ１，ＬＤＨ２和ＬＤＨ４组成；（２）ＲＢＣ－ＬＤＨ同工酶由ＬＤＨ１，ＬＤＨ２和ＬＤＨ３组成，其活性强于     

牦牛卵泡液LDH同工酶及外源性激素对其表达模式的影响

采用比色法和不连续缓冲系统的聚丙烯酰胺凝胶电泳（ＰＡＧＥ）技术对未孕和怀孕初期牦牛卵泡液乳酸脱氢酶（ＬＤＨ）活性及同工酶进行了测定，并分析了促卵泡素３号（ＬＲＨ－Ａ３）、人绒毛膜促性腺激素（ｈＣＧ）和孕马血清促性腺激素（ＰＭＳＧ）对其表达模式的影响。结果表明，未孕和怀孕初期牦牛卵泡液ＬＤＨ活性分别为２００．４±３１．１，２１５．３±８６．６μｍｏｌ·ｓ－１·Ｌ－１，同工酶有５条谱带。３种外源性激素均使牦牛卵泡液ＬＤＨ活性显著升高（Ｐ＜０．０１），ＬＤＨ同工酶带型分布发生变化，降低了ＬＤＨ２／ＬＤＨ１比值，改变了同工酶中Ａ和Ｂ亚基所占比例     

蛭形巨吻棘头虫的LDH同工酶

用聚丙烯酰胺凝胶电泳对蛭形巨吻棘头虫（Ｍａｃｒａｃａｎｔｈｏｒｈｙｎｃｈｕｓｈｉｒｕｄｉｎａｃｅｕｓ）雄虫、雌虫体液和宿主（猪）血清的ＬＤＨ同工酶的酶谱、相对迁移率和相对活性等进行比较表明，雄虫有３条酶带和雌虫有２条酶带显示出很强的特异性；不同浓度的抑制剂（硝硫氰醚）对雄虫和雌虫特有的ＬＤＨ同工酶酶带活性具有不同程度的抑制作用，尤其是雌虫第２带对抑制剂最敏感。蛭形巨吻棘头虫体液具有特异的且不同于宿主（猪）血清ＬＤＨ１～５的ＬＤＨ同工酶酶带，特有的ＬＤＨ同工酶可能由４条不同的区带组成。     

藏獒血液乳酸脱氢酶同工酶酶谱的电泳研究

采用聚丙烯酰胺凝胶电泳法对２１条藏獒的红细胞乳酸脱氢酶（ＲＢＣ－ＬＤＨ）和血清到脱氢酶（Ｓ－ＬＤＨ）同工酶酶谱及其多态性进行研究。结果发现：（１）ＲＢＣ－ＬＤＨ同工酶共有３条区带,呈现两种电泳表型,１１条藏契为ＬＤＨ２Ａ型（５７．８９％）,８条藏獒为ＬＤＨ２ａ（４２．１１％）,（２）Ｓ－ＬＤＨ共有５条区带,呈现两种电泳表型,２条藏獒为ＬＤＨ１Ａ（９．５２％）,１９条藏獒为ＬＤＨ１ａ（９０．４８％）     

大通家牦牛和含野血牦牛的红细胞乳酸脱氢酶同工酶酶谱

采用聚丙烯酰胺凝胶生趣平板电泳地对大通家牦牛，１／４野血牦牛，１／２野血牦牛和３／４野血牦牛的红细胞乳酸脱氢酶同工酶酶谱进行了研究。结果发现，在被检的４个群体中，绝大多数牦牛只有ＲＢＣ－ＬＤＬ１同工酶，部分牦牛逊有ＲＢＣ－ＬＤＨ２和ＲＢＣ－ＬＤＨ３同工酶。     

马立克氏病乌黑鸡群血清学研究

对４月龄感染了马立克氏病的２月龄－６月龄乌黑鸡群血清蛋白及血清酶的研究结果表明;２月龄～３月龄健康乌黑鸡如血清蛋白升高极显著（Ｐ〈０．０１）,血清ＡＬＰ活性下降显著（Ｐ〈０．０５）,ＡＭＳ活性下降不明显（Ｐ〉０．０５）,ＣＨＥ活性升高不明显（Ｐ〉０．０５）,４月龄感染ＭＤ后,血清蛋白显著降低（Ｐ〈０．０５）,而后,白脓月回升,变化均不明显（Ｐ〉０．０５）,白蛋白在５月龄再次极显著降低（Ｐ〈０．０５     

山东青山羊乳酸脱氢酶同工酶电泳的研究

用琼脂糖凝胶电泳技术，在山东青山羊精子ＬＤＨ酶谱中发现了一条新的酶带，称ＬＤＨ－Ｘ酶带，它位置在ＬＤＨ－４与ＬＤＨ－５之间。本文还对山东青山羊血清ＬＤＨ同工酶谱型做了测定，结果表明，青山羊血清ＬＤＨ同工酶谱的特点是ＬＤＨ－１＞ＬＤＨ－３＞ＬＤＨ－２＞ＬＤＨ－５＞ＬＤＨ－４，与其它山羊有一定的差异。     

兔狲血液蛋白质和酶的电泳分析

采用聚丙烯酰胺凝胶电泳法对２只兔狲的４种血液蛋白质表型和３种血液同工酶酶谱进行研究。结果发现：①被检兔狲的血红蛋白（ＨＢ），白蛋白（ＡＬＢ）和后白蛋白（Ｐａ）分别呈单一的ＨＢＡＢ，ＡＬＢＡＡ和ＰａＡＡ型，运铁蛋白（ＴＦ）有ＴＦＡＢ和ＴＦＢＢ两种表型；②血清碱性磷酸酶（ＡＬＰ）由ＡＬＰＡ，ＡＬＰＢ，ＡＬＰＣ和ＡＬＰＤ四种同工酶组成；③血清酯酶（ＥＳ）由ＥＳＩ和ＥＳＩＶ两种同工酶共７条区带组成；④血清乳酸脱氢酶（Ｓ－ＬＤＨ）由ＬＤＨ１，ＬＤＨ２，ＬＤＨ３，ＬＤＨ４和ＬＤＨ５五种同工酶组成，红细胞内只有前四种ＬＤＨ同工酶。     

不同月龄鸵鸟38项血液生化参数比较

比较了 １～３ 月龄（ ｎ ＝ １５）和 ４～６ 月龄（ ｎ ＝ １５）鸵鸟血液的 ３８ 项生化参数。结果，１～３ 月龄和 ４～６ 月龄鸵鸟之间血清 Ｃ Ｋ、 Ｔ Ｇ、 Ｂ Ｕ Ｎ、 Ｂ Ｕ Ｎ／ Ｃ Ｒ Ｅ、 Ｕ Ａ、 Ｍ ｇ、 Ｐ含量差异极显著（ Ｐ ＜ ００１）；血清 Ｔ Ｂ Ａ、 Ｌ Ｄ Ｌ Ｃ、 Ｃ Ｈ Ｏ、 Ｔ Ｔ Ｔ、 Ｎａ、 Ｋ 含量的组间差异显著（ Ｐ ＜ ００５）；其他指标 ２ 组间差异不显著。该结果提示，不同月龄鸵鸟的营养物质代谢过程存在一定差异。     

Growth and Development of Testes in Domesticated and Hybrid(wild × domesticated)Yak Bulls

Testicular tissue was studied in domesticated yak and in wild yak × domesticated yak(F1 and F2)bulls at 6,12,18 and 24 months of age by stereology for quautitative histology and by comparative studies on the lactate dehydrogenase(LDH)isoenzyme spectrum, LDH activity and the percentage content of LDH isoenzymes in testes. The results indicated that all three types of yak were similar in both the characteristics of ultrastructure of testicular tissue and the degree of germ cell development. The process of spermatogenesis was initiated and sperm were produced for the first time at the age of 12months in all three yak types. All yak types exhibited similar age-related, increases in weight of testes,volume density of both the seminiferous tubules and seminiferous epithelium and height of seminiferous epithelium but all these indexes were slightly affected by season. The number of LDH isoenzyme bands after electrophoresis varied with age as follows: four bands were present at 6 months of age, five bands at 12 and 18 months and six bands at 24 months of age. The sixth band was LDH -x ,which appeared between the LDH4 and LDH5 band. These results indicated that yak bulls reach sexual maturity at 24months of age and that puberty is not affected by infusion of wild strains.     

Morphological Observation on the Test and Epididymis in Hybrid of Wild and Domestic Yaks During the Growth Periods

This paper was carried out on the morphological character of the test and epididymis of three types of yak bull age 6,12,18,24 months old (1/2 wild yak, cross 1/2 wild yak and domestic yak). The results showed that the weight and size of testes and epidiymis was increased with the age which was hardly affected by imbalance of food supply during the cold and warm seasons. The development were similar for three kinds of yak at the same age(P＞0.05).     

Study on The Isozyme of Wild Yak

We have adopted PAGE method to isolate lactic dehydrogenase(LDH)and α-amylase(α-Am)from wild yak's serum and measure the physical-chemical properties of the two components respectively. The results showed that the yak LDH isozyme distributed into five bands. The sequence according to their activity reads as LDH1 ＞ LDH2＞ LDH3 ＞ LDH4 ＞ LDH5. The subunit B is relatively occupied a dominant position. It got a wide operating pH range, high thermal stability,well denature resistance and a fairly large adaptability threshold on the physical-chemical change of surrounding environment. α-Am isozyme distributed into seven bands and also got a wide operating pH range. α-Am isozyme is high sensitivity to environmental temperature.     

Quantitative Histology Studies on the Test in Hybrid Bull of Wild and Domestic Yak

The testicular tissue of three types of yak bull (1/2 wild yak,cross 1/2 wild yak and domestic yak) were studied quantitatively at 6,12,18 and 24 months old. The results showed that the average values changed from breed to breed at the same age. But there were no significant difference. The volume density and the height of seminiferous tubule and epithelium increased with the age and testicular weight. The capacity rate of the testicular seminiferous tubule in three types were 78.71% ,75.78% and 78.58% respectively, which nearly reached the level of mature bull.     

犏牛PPP1R11基因的克隆及在睾丸中的表达规律研究

旨在克隆犏牛蛋白磷酸酶1调节亚基11(protein phosphatase 1 regulatory subunit 11,PPP1R11)基因,分析其在不同发育阶段睾丸中的表达与定位,为解析其在雄性生殖中的功能机制提供理论依据.本研究采集成年犏牛睾丸、附睾、心、肝、脾、肺、肾、大肠、小肠、胃、肌肉和脂肪组织(n=3...     

The Expression of Epidermal Growth Factor (EGF) and its Receptor (EGFR) During Post‐Natal Testes Development in the Yak

Growth factors play critical role in cell proliferation, regulate tissue differentiation and modulate organogenesis. Several growth factors have been identified in the testes of various mammalian species in last few years. In present investigation, the objective was to determine the expression of epidermal growth factor (EGF) and the epidermal growth factor receptor (EGFR) in yak testicular tissue by relative quantitative real time polymerase chain reaction (RT‐PCR), Western blot (WB) and immunohistochemistry (IHC) from mRNA and protein levels. The testicular tissues were collected from male yak at 6 and 24 months old. Results of RT‐PCR and WB showed that the expression quantity of EGF and EGFR at 24 months of age was higher than at 6 months, and the increase rate of EGFR on mRNA and protein levels was higher than the increase rate EGF during post‐natal testes development. Positive staining for EGF and EGFR was very low and mainly localized to Leydig cells testes at 6 months of age with immunohistochemistry, and seminiferous tubules were not observed. At 24 month of age, both the EGF and EGFR could be detected in Leydig cells, peritubular myoid cells, sertoli cells and germ cells of the yak testes. However, EGF and EGFR were localized to preferential adluminal compartment and basal compartment in the seminiferous tubules, respectively. In conclusion, the findings in present studies suggest that EGF and EGFR as important paracrine and/or autocrine regulators in yak testes development and spermatogenesis.     

导入野血牦牛群体的遗传变异和基因分化

采用聚丙烯酰胺凝胶电泳法和火焰光度法对家牦牛和３个含野血牦牛群体ＴＦ,ＨＢ,Ｈｐ,ＫＥ,ＡＭＹ１和ＲＢＣＬＤＨ１６个基因座多态性的研究表明：家牦牛与３个含野血牦牛群体具有同样的多态性特征,但群体的遗传变异随野牦牛血比例的增多而减少。含野血牦牛群体之间的亲缘关系大于家牦牛,再大于野牦牛,但总群体所发生的基因分化程度很小（２７４８％）。     

肃南县牦牛导入青海大通牛场野牦牛血液复壮效果调查

[目的]通过引进野血种公牦牛改良当地牦牛可有效遏止牦牛退化,进一步巩固和改善我县牦牛品种质量,恢复其原有的优良生产性能和品种特征,提高经济效益.[方法]本文以野牦牛和家牦牛杂交后代与家牦牛后代为研究对象,分别测定了杂种牦牛和家牦牛的初生、6月龄、18月龄体尺、体重.[结果]显示,在同等条件下,1/4野血牦牛初生、6月龄、18月龄体尺、体重均显著高于同龄家牦牛,经t检验差异极显著（P＜0.01）.[结论]用野血牦牛改良家牦牛,是目前更新复壮家牦牛群体的有效途径之一,能获得很强的杂种优势,对提高经济效益,增加农牧民收入,保护生态环境都具有十分重要的意义.     

野牦牛及青海地方牦牛品种全线粒体基因组母系遗传多样性、分化及系统发育分析

旨在从分子水平上探究野牦牛及青海地方牦牛品种的母系遗传多样性、群体遗传结构、亲缘关系和遗传背景。本研究在测定青海省4个地方牦牛品种(即青海高原、环湖、雪多和玉树牦牛)22条全线粒体基因组(Mitogenome)序列的基础上,从GenBank下载了已公布的野牦牛及上述4个地方牦牛品种的142条相应序列,使用BioEdit 7.2.5、Arlequin 3.11和Network 10.1等软件对共计164条线粒体基因组序列进行综合分析。结果显示：1)根据序列间核苷酸变异共确定了115种单倍型,其中野牦牛和青海地方牦牛品种分别拥有22种和93种单倍型;在野牦牛和青海高原、环湖、雪多、玉树牦牛中分别检测到22、26、18、23、19种特有的单倍型。遗传多样性分析显示,野牦牛单倍型多样度最高(0.992 8±0.014 4),且高于4个青海地方牦牛品种的单倍型多样度(0.973 1±0.007 7);4个青海地方牦牛品种单倍型多样度大小依次为：雪多牦牛(0.988 5±0.012 6)、玉树牦牛(0.975 8±0.018 7)、青海高原牦牛(0.973 0±0.016 6)和环湖牦牛(0.939 3±0.027 8)。2)野牦牛与环湖牦牛之间的固定分化指数值(F_ST值)最大(0.041 2),分化程度最高,而与玉树牦牛间的F_ST值最小(-0.008 8),分化程度最低。青海4个地方牦牛品种中,雪多牦牛与青海高原牦牛之间F_ST值最大(0.035 8),分化程度最高,而雪多牦牛与环湖牦牛间F_ST值最小(0.011 2),分化程度最低。3)聚类分析显示,4个青海地方牦牛品种各自为1类,存在明显的母系遗传差异。相比而言,环湖牦牛和雪多牦牛聚类较近,青海高原牦牛和玉树牦牛聚类较近,而野牦牛与玉树牦牛聚类关系更近,各品种(群体)间的聚类结果与其分化程度、地理分布一致。4)系统发育分析表明,115种单倍型分布在3个大的母系遗传分支(即Mt-Ⅰ、Mt-Ⅱ和Mt-Ⅲ),其中Mt-Ⅰ支系所占比例为72.17%,由A、B、E和F 4种单倍型组构成;Mt-Ⅱ支系包括C、D和H 3种单倍型组,占26.09%;而Mt-Ⅲ支系只包含G单倍型组,由雪多牦牛和野牦牛所拥有,所占比例为1.74%,提示牦牛有3个母系起源。综上所述,野牦牛和青海4个地方牦牛品种均具有丰富的母系遗传多样性,其多样性水平由高到低依次为野牦牛、雪多牦牛、玉树牦牛、青海高原牦牛和环湖牦牛。青海4个地方牦牛品种间及与野牦牛间的遗传分化程度均较弱,但各自拥有特有的母系遗传信息,存在明显的母系遗传差异。野牦牛和青海家牦牛品种由3个母系支系组成,推测牦牛有3个母系起源。     

犏牛和牦牛ESCO2基因的序列特征及其在睾丸中的表达对比分析

旨在克隆犏牛和牦牛姐妹染色单体内聚建立蛋白2（establishment of sister chromatid cohesion N-acetyltransferase 2,ESCO2）基因,并分析其在不同发育阶段睾丸中的表达与定位,为进一步解析ESCO2在减数分裂过程中的作用机制提供理论依据。本研究以健康雄性犏牛及牦牛为试验动物,根据年龄分为胎牛组（5~6月龄）、幼年组（1~2岁）和成年组（3~4岁）,每组各3头。通过RT-PCR技术克隆犏牛和牦牛ESCO2基因并进行生物信息学分析,采用实时荧光定量PCR （quantitative real-time PCR,qRT-PCR）检测ESCO2基因在犏牛不同组织中的表达谱,比较分析ESCO2在犏牛和牦牛不同时期睾丸中的表达规律,利用免疫组织化学（immunohistochemistry,IHC）技术检测ESCO2蛋白的细胞定位和表达差异。结果显示,犏牛ESCO2基因（GenBank登录号：MW198470） CDS区为1 833 bp,编码610个氨基酸,与牦牛相比,犏牛ESCO2序列第301~319位多19个氨基酸,另有3个氨基酸突变;犏牛ESCO2蛋白序列与黄牛的同源性高于其他哺乳动物;ESCO2可能与SMC3、SMC1A、PDS5A、PDS5B、STAG2等蛋白相互作用,互作蛋白功能与姐妹染色单体凝聚、减数分裂细胞周期、DNA修复、细胞分裂和染色体重构等生物学过程相关。ESCO2在犏牛各组织中均有表达,但在睾丸中的相对表达水平显著高于其它组织（P<0.05）;在犏牛睾丸中的表达随年龄增长呈上升趋势,幼年和成年时期犏牛睾丸中ESCO2的表达显著低于同时期牦牛（P<0.05）;IHC染色结果发现,雄性犏牛减数分裂阻滞于初级精母细胞,ESCO2蛋白在犏牛初级精母细胞中无表达并与牦牛存在差异。本研究结果表明,犏牛与牦牛的ESCO2基因、蛋白序列差异较大,且在睾丸发育过程中的表达模式差异显著,这可能是引起雄性犏牛减数分裂阻滞及不育的原因之一,其具体作用机制有待进一步研究。