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为了探究磷脂酶patatin样域包含蛋白8(patatin-like phospholipase domain containing 8,PNPLA8)在水牛乳腺脂质代谢中的作用,试验根据GenBank中公布的奶牛PNPLA8基因序列(登录号:XM_005205444.4)设计引物,应用PCR扩增并克隆水牛PNPLA8基因编码区(CDS),应用生物信息学软件分析序列及蛋白质结构;抽提水牛不同组织及不同泌乳期乳腺组织RNA,利用实时荧光定量PCR检测PNPLA8基因在不同组织间和不同泌乳期的表达;利用不同浓度的催乳素处理水牛乳腺上皮细胞,通过定量检测催乳素对PNPLA8基因表达的影响。结果显示,水牛PNPLA8基因CDS长2 355 bp,编码784个氨基酸,与牦牛、山羊等PNPLA8基因具有较高的同源性;PNPLA8基因在所检测的水牛11个组织中有不同水平的表达,在肺脏和乳腺中表达量相对较高,在脂肪和肌肉组织中表达量较低;在整个泌乳期内PNPLA8基因的表达呈现"低-高-低"趋势;催乳素处理水牛乳腺上皮细胞结果显示,随着催乳素浓度升高,PNPLA8基因表达量逐渐下降。本研究成功克隆了水牛PNPLA8基因,并发现PNPLA8基因是参与乳腺泌乳的一个功能基因,为进一步研究PNPLA8基因在水牛乳腺中的功能奠定基础。 相似文献
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《黑龙江畜牧兽医》2010,(21)
为了对奶牛乳腺肥大细胞的组织化学特点和分布特点进行研究,试验对取自21头发育各期奶牛的乳腺样品,应用免疫组织化学方法(SP)观察奶牛乳腺肥大细胞5-羟色胺(5-HT)和类胰蛋白酶的表达。结果表明:5-羟色胺和类胰蛋白酶在奶牛乳腺肥大细胞中表达,肥大细胞数量在各期有显著差异,泌乳期的乳腺肥大细胞数量明显减少(P0.05),分娩后60天奶牛乳腺肥大细胞的数量最少;5-羟色胺在不同发育时期奶牛乳腺组织中的表达量少于类胰蛋白酶在不同发育时期奶牛乳腺组织中的表达量。说明肥大细胞的数量随生理周期的改变而增减的趋势可能与乳腺的发育、乳腺局部免疫状态及内分泌调节有密切关系。 相似文献
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为了研究昆明小鼠乳腺发育4个时期即青春期、妊娠期、泌乳期和退化期的同源异型盒基因(Hex)蛋白表达规律及组织定位,试验采用免疫组织化学法进行检测。结果表明:昆明小鼠乳腺组织结构呈周期性变化,青春期至妊娠期Hex蛋白逐渐增加,泌乳期达到高峰,退化期逐渐下降至类似青春期水平;Hex蛋白主要定位于细胞质中,泌乳期在细胞核中也有少部分出现;昆明小鼠乳腺发育的4个阶段均有Hex蛋白表达,且与乳腺周期性的变化一致。说明Hex蛋白参与乳腺的发育和泌乳调控过程。 相似文献
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《中国兽医学报》2016,(6)
与奶牛乳脂相关的基因在奶牛乳腺组织中表达量的多少对奶牛的乳脂率起到关键的调控作用。C4BPA(complement component 4binding protein,alpha)基因是通过对高乳脂奶牛及低乳脂奶牛的乳腺组织进行转录组测序选择出的差异基因。为了研究C4BPA基因在奶牛乳脂合成代谢过程中的作用,本试验以中国荷斯坦奶牛为研究对象,利用乳成分分析仪检测15头中国荷斯坦奶牛的乳脂率,挑选出高乳脂奶牛与低乳脂奶牛各3头(P0.05),提取乳腺组织总RNA,反转录成cDNA,进行荧光定量PCR,检测该基因在奶牛乳腺组织中的表达量,并利用SPSS软件对不同奶牛乳腺组织中C4BPA基因的表达量进行差异分析。结果表明,C4BPA基因在不同奶牛乳腺组织中均有表达,且在高乳脂奶牛及低乳脂奶牛的乳腺组织中表达量差异显著(P0.05)。本试验为进一步对C4BPA基因生物学功能和作用机制的研究奠定基础。 相似文献
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《中国畜牧兽医》2016,(9)
γ-氨基丁酸(GABA)是大脑中一种重要的抑制性递质,除与兴奋的传递有关外,还在内分泌组织中发挥着重要的作用。本研究对水牛GABA A型受体的π亚基基因(GABRP)CDS序列进行了克隆,并对其进行了生物信息学分析,同时应用实时荧光定量PCR和免疫组化技术分析了GABRP基因在水牛乳腺组织中的表达情况。结果表明,应用RT-PCR技术成功获得了广西本地水牛GABRP基因序列,长为1 401bp;BLAST比对结果显示,广西本地水牛的GABRP核苷酸序列与和河流型水牛的同源性为99%,GABRP在不同物种中具有较高的保守性;GABRP蛋白主要在水牛乳腺组织的腺泡上皮细胞中表达,GABRP基因在泌乳期及非泌乳期水牛乳腺组织中都有表达,且非泌乳期GABRP基因的表达显著高于泌乳期(P0.05)。试验结果为进一步研究GABRP基因在水牛乳腺发育过程中的功能奠定了基础。 相似文献
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为探讨脾源性酪氨酸激酶(spleen tyrosine kinase,SYK)的表达与奶牛乳腺发育和泌乳功能之间的关系,试验采用Western blotting和激光共聚焦显微技术对泌乳期高乳品质、低乳品质及干乳期的中国荷斯坦奶牛乳腺组织中SYK的表达含量和表达部位的变化进行研究。结果表明,干乳期奶牛乳腺组织中SYK的表达显著高于泌乳期奶牛乳腺组织(P<0.05),泌乳期高乳品质、低乳品质奶牛乳腺组织中SYK的表达差异不显著(P>0.05);在干乳期SYK主要在乳腺导管上皮细胞的胞质中表达,而在泌乳期SYK在腺泡上皮细胞中表达。结果提示SYK是乳腺上皮细胞增殖与分化的调节因子,主要参与干乳期乳腺组织的重建过程。 相似文献
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《畜牧兽医科技信息》2015,(8)
试验选取青春期、妊娠期、泌乳期(分产高品质奶与产低品质奶)与退化期等几个时期的奶牛乳腺组织,提取RNA,实时荧光定量PCR检测各组织中I型和II型肽转运蛋白的表达情况。结果表明,I型肽转运蛋白的表达在青春期较低,在妊娠期与泌乳期显著上升,至泌乳期达到顶峰,在退化期下降至青春期水平;II型肽转运蛋白在各个时期表达无明显变化;在泌乳期高品质乳组织中,I型肽转运蛋白的表达显著高于低品质乳组织,且在这两组样品中,I型肽转运蛋白的表达均显著高于II型肽转运蛋白的表达。结果提示,在奶牛泌乳过程中,I型肽转运蛋白与泌乳的调节有显著的正相关性,而II型肽转运蛋白与泌乳的调节无明显关系。 相似文献
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水牛泌乳期与非泌乳期乳腺组织差异miRNAs的表达模式鉴定及分析 总被引:1,自引:0,他引:1
为了探求水牛泌乳和非泌乳期关键调控差异表达miRNAs及其表达模式,试验结合Solexa高通量测序和生物信息学分析结果筛选出18个水牛乳腺组织在泌乳期和非泌乳期差异表达的miRNAs,设计、筛选可行的miRNAs反转录及实时荧光定量PCR引物,进行实时荧光定量PCR验证差异表达miRNAs的表达模式。结果发现,miRNA的差异表达模式与测序结果基本一致。泌乳期miRNA-103、miRNA-125a、miRNA-30a-5p和miRNA-148a的表达量均显著高于非泌乳期(P<0.05);miRNA-29a在非泌乳期表达量显著高于泌乳期(P<0.05);miRNA-141、miRNA-125b和miRNA-497在泌乳期表达量均高于非泌乳期,但差异不显著(P>0.05);低丰度的miRNA-490和miRNA-592在泌乳期表达量均显著高于非泌乳期(P<0.05)。Novel-123b在泌乳期和非泌乳期差异不显著(P>0.05),Novel-57在泌乳期比非泌乳期高29.79倍(P<0.01)。本试验结果为后续水牛乳腺研究提供了16个miRNA的可用特异性反转录引物和实时荧光定量PCR引物,miRNA-103、miRNA-125a、miRNA-30a-5p、miRNA-148a、miRNA-29a和Novel-57 6个水牛差异表达miRNAs值得进一步研究。 相似文献
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为探究小鼠钠碘转运体(Na+/I- symporter,NIS)蛋白的结构、功能及其在乳腺组织的表达特征,本研究扩增小鼠乳腺NIS基因编码区,测序鉴定后采用结构预测软件对NIS蛋白进行生物信息学分析,同时采用Western blotting、免疫组织化学技术和半定量PCR技术对NIS基因在不同乳腺发育时期的转录和表达水平进行分析。结果显示,小鼠NIS基因编码区全长1 857 bp,与人NIS的同源性为80.40%,保守性较强。NIS蛋白含有618个氨基酸残基,分子质量约为65.7 ku,分子式为C2996H4701N749O833S32,理论等电点为7.44,半衰期为30 h,消光系数为102 510,不稳定系数为29.31,脂溶系数为105.91,富含疏水性氨基酸,为稳定的疏水蛋白。NIS蛋白二级结构含有α-螺旋(41.42%)、延伸链(18.28%)、β-转角(4.05%)和无规则卷曲(36.25%),三级结构与之相一致。Western blotting结果显示,4个时期均出现特异性杂交带,大小为65.7 ku,与预期结果相符。免疫组织化学结果表明,NIS蛋白在小鼠的青春期、妊娠期到泌乳期的乳腺组织中表达量逐渐增加,在泌乳期达到高峰,存在显著差异(P<0.05),在退化期逐渐恢复到类似青春期状态;NIS基因mRNA的变化规律与蛋白表达一致。本研究结果为进一步探究NIS蛋白在乳腺发育和泌乳调控中的作用提供了参考依据。 相似文献
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为获得奶山羊CIDEa基因序列并检测其在乳腺组织中的表达,采用RT-PCR技术从奶山羊乳腺组织扩增CIDEa基因序列,利用生物信息学软件对其进行预测分析,并采用荧光定量PCR检测CIDEa在干奶期和不同泌乳时期乳腺组织中的表达。结果表明:通过克隆测序得到奶山羊CIDEa基因CDS区660 bp,编码219个氨基酸,与GenBank公布的绵羊、牛、人和猪的核苷酸序列同源性分别为99%、96%、85%、85%。奶山羊CIDEa蛋白属于碱性、不稳定亲水蛋白,不存在跨膜结构和信号肽;蛋白主要定位在细胞质、线粒体和细胞核。CIDEa蛋白结构主要由α螺旋、β折叠和不规则卷曲组成。奶山羊CIDEa在泌乳前期、盛期和中期乳腺组织中表达量均显著高于干奶期(P<0.05)。说明CIDEa可能参与奶山羊乳腺的泌乳过程,为进一步研究其在奶山羊乳腺组织中的功能及对乳品质的调控作用奠定基础。 相似文献
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MA Xiaoya PANG Chunying LU Xingrong DUAN Anqin LIANG Shasha LIANG Yanyin DENG Tingxian LIANG Xianwei 《中国畜牧兽医》2007,47(9):2715-2723
In order to investigate the role of patatin-like phospholipase domain-containing 8(PNPLA8) in lipid metabolism of mammary gland in buffalo,the coding region (CDS) was amplified and cloned by PCR based on the sequence of Bos taurus PNPLA8 gene in GenBank (accession No.:XM_005205444.4) were analyzed using bioinformatics software.Total RNA was extracted from different tissues of buffalo and mammary glands,which were harvested from different lactating buffaloes.The expression of PNPLA8 gene mRNA in different tissues and different lactation period was detected by Real-time quantitative PCR.For buffalo mammary epithelial cell treatment,different concentrations of prolactin were used and the effect of prolactin on the expression of PNPLA8 gene was detected by Real-time quantitative PCR.The results showed that the length of the PNPLA8 gene CDS was 2 355 bp,encoding 784 amino acids.The sequence showed high homology with Bos mutes and Capra hircus.PNPLA8 gene was expressed at different levels in 11 tissues examined,with a relatively high level in the lung and mammary tissues while the low level in the fat and muscle tissues.The expression abundance of the PNPLA8 gene was variable during lactation and showed a trend of "low-high-low".Prolactin treatment showed that the expression of PNPLA8 gene decreased with the increase of prolactin concentration.In this study,PNPLA8 gene of buffalo was successfully cloned,and the expression of PNPLA8 gene in different tissues and the lactation period was analyzed.Herein,the effect of prolactin on the expression of PNPLA8 gene was studied that laid a foundation for further research on PNPLA8 gene of mammary gland in buffalo. 相似文献
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Onda K Matsuki N Ono K Wada Y 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2006,68(1):9-13
The expression of parathyroid hormone-related protein (PTHrP) mRNA was examined in mammary gland with or without lactation, and during periparturient period in a Holstein cow and a Jersey cow. In the lactating mammary gland, PTHrP was detected in alveolar epithelial cells and the lumen by immunohistochemical analysis. The relative expression levels of PTHrP mRNA in mammary gland from lactating cows were significantly higher than those from non-lactating cows (P<0.05). During periparturient period, relative PTHrP mRNA level was remarkably low before the parturition in a Jersey and a Holstein cow, however, both levels were gradually increased and reached a peak level at 5-6 weeks after the parturition. In addition, the peak level in a Jersey cow was approximately 3-fold higher than that in a Holstein cow. From these results, PTHrP was synthesized and secreted in alveolar epithelial cells in mammary gland and increased subsequently with the lactation, suggesting a possible mechanism for the regulation of local calcium homeostasis. 相似文献
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本研究旨在对山羊乙酰辅酶A合成酶2(acetyl-CoA synthetase 2,ACSS2)基因进行克隆和生物信息学分析,并检测其在山羊不同泌乳时期乳腺组织中的表达量变化。以山羊乳腺组织RNA为模板,采用RT-PCR方法扩增并克隆山羊ACSS2基因完整CDS区序列,对测序结果进行生物信息学分析,并对ACSS2基因在山羊不同泌乳时期乳腺组织中的表达量进行分析。结果显示,山羊ACSS2基因CDS区序列长2 106 bp,编码701个氨基酸;山羊ACSS2基因与牛、马、人、犬、猪、小鼠和鸡的同源性分别为97.8%、92.0%、91.3%、91.3%、91.1%、88.1%和73.3%。蛋白理化性质分析结果表明,ACSS2蛋白分子质量为78.72 ku,理论等电点为6.03,属于酸性蛋白;跨膜结构和信号肽分析表明,ACSS2蛋白不含跨膜结构和信号肽;结构域分析表明,该蛋白含有1个乙酰辅酶A合成酶N端结构域。亚细胞定位分析结果表明,该蛋白主要分布在内质网(44.4%)、线粒体(33.3%)、细胞质(11.1%)和细胞核(11.1%)中。蛋白质结构预测发现ACSS2蛋白含有α-螺旋(29.10%)、延伸链(21.54%)、β-转角(9.84%)及无规则卷曲(39.52%)。实时荧光定量PCR分析结果表明,ACSS2基因在不同泌乳时期均有表达,其中在泌乳中期表达量最高,在干奶期表达量最低。本试验结果为进一步研究山羊ACSS2基因在脂质代谢过程中的功能及转录调控机制提供了参考。 相似文献
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HAO Wen-yan CHEN Pei-fang DAI Xiao-li WANG Ping WU Da-ping SHI De-shun LI Xiang-ping 《中国畜牧兽医》2016,43(9):2412-2417
GABA is an important inhibitory neurotransmitter in the brain,beside the function of transmissing information,which plays an important role in endocrine tissues.In this study,the CDS of buffalo GABRP gene was cloned and its biological information was analyzed,the expression of GABRP in buffalo mammary tissue was also examined by Real-time quantitative PCR and immunohistochemistry.The RT-PCR results showed that GABRP CDS fragment was obtained successfully from Guangxi local buffalo,which shared 99% with that of river buffalo,its length was 1 401 bp.The phylogenetic tree showed that GABRP had high conservation in different species.The immunohistochemistry results showed that buffalo GABRP protein located at the alveolar epithelial cells of buffalo mammary.The expression of GABRP gene in non-lactation buffalo mammary was significantly higher than that of lactation tissue (P<0.05).The results laid a foundation for further study the function of GABRP gene in the development of buffalo mammary gland. 相似文献
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LIN Lin FAN Hui-quan XING Wei-nan YANG Yang HOU Xiao-ming ZHANG Chang-shun LIN Ye 《中国畜牧兽医》2015,42(5):1240-1244
To investigate the relationship between the expression of SYK and dairy cow mammary gland development and lactation, the expression of SYK in lactating dairy cow mammary gland with high or low quality milk and dry period Holstein dairy cow mammary gland was detected by Western blotting and laser confocal microscope.The results showed that SYK expression in dry period mammary gland was significant higher than that in lactating mammary gland (P<0.05).There was no SYK differential expression detected between lactating mammary gland with high quality milk and low quality milk (P>0.05).SYK was mainly located in the cytoplasm of ductal epithelial cells in dry period mammary gland.In lactating mammary gland, SYK was existed in acinar epithelial cells.All these results revealed that SYK was a regulator in mammary epithelial cell proliferation and differentiation.It participated in mammary gland reconstitution in dry period. 相似文献