鹌鹑黑素皮质素受体-4基因的克隆与序列分析

黑素皮质素受体-4(MC4R)是G蛋白偶联受体超家族的一个成员,在人、鼠和鸡的体重、能量稳态和采食量的调控中具有重要作用。实验根据GenBank中鸡MC4R基因全序列设计9对引物,对北京白羽蛋用鹌鹑和法国沙维玛特肉用鹌鹑MC4R基因序列进行PCR扩增及克隆测序,获得的序列长度分别为2154bp和2152bp。结果表明:所测的两段序列经GenBank中的Blast分析与鸡MC4R基因同源性均为97%;在MC4R基因核苷酸水平上,北京白羽蛋用鹌鹑与法国沙维玛特肉用鹌鹑的同源性为99.11%;在MC4R)基因氨基酸水平上两品种鹌鹑同源性也很高。     

绒山羊黑素皮质素受体4（MC4R）基因的克隆与序列分析

绒山羊是绒肉兼用型动物,国内外对绒山羊绒用性能研究广泛,但对肉用性能研究相对较少。黑素皮质素受体4（melanocortin 4receptor,MC4R）主要分布在脑组织和肾上腺髓质及皮肤中,该基因突变会引起动物的体质量、能量稳态及采食量的改变。本研究采用PCR及RT-PCR等技术克隆得到了绒山羊MC4R基因的DNA及cD-NA片段,对比发现MC4R无内含子,长924bp,编码304个氨基酸,具有7个跨膜结构域。同源性比较显示：与绵羊、牛、马、猪、人、小鼠、兔、犬等同源性均高于85%,与绵羊同源性最高,达98%;与系统进化树分析结果一致。与其他哺乳动物相比,9处突变频率较高,分别位于93、152、160、162、165、182、213、221、230,且这些突变大多位于MC4R蛋白跨膜结构域及拓扑结构域,该研究对改良哺乳动物繁育性状提供了依据,对提高绒山羊肉用市场价值的研究发挥重要作用。但该突变是否与绒山羊特殊的毛肉用兼用性能有关,还有待进一步研究。     

水貂黑素皮质素受体-4基因部分序列的克隆测序

对水貂黑素皮质素受体-4(MC4R)基因进行了克隆和序列分析,以期为进一步开展水貂MC4R基因与其肥胖性状的相关分析及基因定位等研究提供理论基础。首先采用特定引物对水貂MC4R基因进行PCR扩增、克隆和测序,然后用DNAMAN软件拼接MC4R基因全序列,并进行序列的同源性分析。试验获得水貂MC4R基因序列816 bp。水貂MC4R基因序列与同一种属的水獭的同源性最高,达96%,而与犬、狐、大熊猫、河马、野猪、山羊、人及小鼠这些哺乳动物的同源性在89%～95%之间,另外,与贝加尔湖海豹和加州海狮的同源性也较高,均为94%。本研究成功克隆了水貂的MC4R基因,结果表明,其在物种间具有较高的保守性。     

鸡黑素皮质素受体-4多肽抗体的制备与鉴定

根据GenBank中报道的鸡黑素皮质素受体-4(melanocortin-4 receptor,MC4R)一级结构信息,用TMHMM 2.0和DNAstar软件分析MC4R蛋白的跨膜结构域、二级结构、疏水性、亲水性以及抗原性等,综合考虑抗体设计的其他因素,设计出1段15个氨基酸的多肽.将合成后的多肽与牛血清白蛋白(BSA)偶联,用与BSA偶联的MC4R多肽免疫新西兰兔,3个月后获取血清,亲和纯化出抗MC4R多肽抗体.通过ELISA法检测效价,Western blot和免疫组化法检测抗体特异性.通过该方法得到了高效价与高特异性的鸡MC4R多肽抗体,ELISA法测定其效价可达到1∶128 000,而且该抗体可用于免疫组化,说明所制备的鸡MC4R抗体具有高特异、高效价等特点,将为鸡MC4R基因的功能研究提供物质基础.     

藏猪黑素皮质激素受体-4基因TaqI多态性分析

藏猪是我国青藏高原特有的小型原始地方猪种,文中采用PCR-RFLP技术分析了藏猪黑素皮质激素受体-4(MC4R)基因TaqI酶切位点多态性。结果表明:藏猪MC4R基因TaqI酶切位点优势基因型为11,基因型频率达93.56%,与国内梅山猪、二花脸猪、民猪和金华猪等较接近,与汉普夏、大白、杜洛克等外种猪差别较大。     

猪黑素皮质素受体-4基因在成纤维细胞中的沉默研究

黑素皮质素受体-4(melanocortin receptor-4,MC4R)、黑素皮质素受体-3(melanocortin receptor-3,MC3R)、瘦素(Leptin)和缩胆囊素受体(cholecystokinin A receptor,CCKAR)基因均是与采食量相关的功能基因,为了研究这4个基因之间可能存在的互作关系,本研究采用RNAi方法,将体外培养的成纤维细胞中的MC4R基因沉默,然后采用Real-time PCR方法对Leptin、MC3R和CCKAR基因的表达变化情况进行了研究。结果表明,MC4R被沉默后,Leptin、MC3R和CCKAR的表达水平均没有发生显著变化。     

黑素皮质激素受体1基因与牛的毛色

简述了牛毛色形成的机制,并介绍了牛MC1R基因的定位、突变、多态检测及对毛色的影响。最后对牛毛色形成的调控机制提出了一个假设模型,并对MC1R基因进一步研究提出了一些看法。     

黑素皮质激素受体1基因与牛的毛色

简述了牛毛色形成的机制,并介绍了牛MC1R基因的定位、突变、多态检测及对毛色的影响.最后对牛毛色形成的调控机制提出了一个假设模型,并对MC1R基因进一步研究提出了一些看法.     

比格犬MC3R和MC4R基因多态性与体重相关性的研究

犬体重是重要的经济性状,为探讨犬MC3R和MC4R基因多态性与犬体重性状的关系,本研究利用PCR-R FLP、AS-PCR方法对120只比格犬MC4R基因T101N位点,MC3R基因的A167T位点,G291A位点的不同基因型与体重的关系进行最小二乘分析、结果表明:对于MC4R基因NN基因型的比格犬体重显著高于其他2个基因型(P<0.05),而MC3R基因中TT和AA基因型的比格犬体重则显著高于其他2个基因型(P<0.05)。在MC4R基因型一致时,NNTT和NNAA基因型的犬体重显著高于其他合并基因型(P<0.01),NNTTAA合并基因型的犬体重最高,MMAAGG基因型犬体重最低。实验结果表明,MC3R和MC4R基因可以作为犬体型的候选基因,可筛选MC3R和MC4R合并基因型的犬用于培育不同体重的犬。     

四环素诱导的猪黑素皮质激素受体-4基因真核表达载体的构建

猪黑素皮质激素受体-4(melanocortin-4 receptor,MC4R)基因是影响猪生长肥育的主效基因之一,为了进一步研究MC4R基因的生物学功能,制备高成活率转基因猪,通过采用条件性诱导表达载体系统,PCR方法扩增了猪MC4R基因完整编码区,四环素诱导表达系统(Tet-on)的启动子调控区与转录调节元件。采用双酶切和连接等方法将上述元件连接到pcDNA3.1(+)上,构建了重组质粒pcDNA3.1(+)-rtTA-P_Tight-MC4R。经双酶切和测序鉴定,结果显示片段正向连接且片段全长测序正确。试验成功构建了四环素诱导的pcDNA3.1(+)-rtTA-P_Tight-MC4R真核表达载体, 并能在时间特异性方面调控MC4R基因的表达。     

浅谈犬的视觉

眼睛是动物的重要感觉器官,用于获取外部环境信息,适应周围的环境。对于犬而言,视觉的重要性仅次于嗅觉,犬可以通过视觉接受人的命令,了解所在地点的情况,进而完成各项工作,人类只有通过了解犬的视觉,才能明白犬所能看到的世界是怎样的,从而更好地与犬交流,使它更加出色地工作。在所有动物中,犬的视力大约是中等水平。若和人的视力相比较,大约只有人的20%～40%,这是因为犬的眼球水晶体比较大,较难调节远近,视力范围为20～30米,这也是说犬是“近视眼”的原因。据实验,犬只能看清50米之内静止的物体,但是对移动的物体具有特别的侦视能力,可达…     

黑皮质素4型受体在绵羊下丘脑区域的定位

黑皮质素4型受体(melanocortin 4 receptor,MC4R)作为下丘脑参与能量调控过程中的关键性受体,在动物能量代谢调节中被广泛关注。目前,关于该受体在绵羊下丘脑中分布区域的相关报道较少。针对试验过程中MC4R免疫荧光结果不稳定,分布区域不明显的特点,本研究在改进现有MC4R免疫荧光试验方法基础上,对MC4R在绵羊下丘脑的分布区域进行了观察。结果显示,分别经过4%多聚甲醛长时间固定(6 d),Triton X-100(pH≈8.68)长时间通透(2 d),以及3%过氧化氢和0.2 mol/L甘氨酸+0.3%SDS+20%甲醇的预处理后,可以获得稳定且饱满的MC4R免疫荧光结果。MC4R主要分布在绵羊下丘脑的室旁核(arcuate nucleus,ARC)区域,但在弓状核区域并无类似分布。     

对犬真菌性疾病的探讨

在犬病临床中,犬的真菌性疾病占有相当大的比例。真菌性疾病的感染主要表现为外源性和内源性感染,外源性感染主要是犬的真菌性皮肤病及其他的一些疾病,而内源性感染主要包括组织胞浆菌病、球孢子菌病、芽生菌病、隐球菌病、孢子丝菌病、念珠菌病等。我们常常对犬的外源性的真菌性皮肤病能及早地做出诊断和治疗,但往往容易对犬的真菌性内源性感染忽略或意识不到。     

谈犬的屠宰检疫

随着人们生活水平的提高,人们的饮食结构也在逐渐发生变化.犬肉由于具有风味独特、营养丰富、健康保健的优点,越来越受广大人们的青睐,食用犬肉安全问题也成为人们关心的话题.目前,世界卫生组织规定的67种B类动物疫病中,犬易患的狂犬病、伪狂犬病、旋毛虫病就包含其中,这三种病也是我国规定的二类动物疫病.犬是杂食动物,活动范围广,接触人和其他动物的机会多,感染人畜共患传染病和寄生虫病的机会相应也多,因而,加强犬的屠宰检疫尤为重要.     

N-ras mutation in a canine lymphoma: short communication

Lymphomas of dogs were investigated by molecular genetic methods. Regions of exon 1 and 2 of the N-ras gene, which harbours the mutation hot spots (codons 12, 13 and 61) were screened. A GGT [symbol: see text] GAT (glycine [symbol: see text] aspartic acid) mutation in codon 13 was present in a multicentric-type lymphoma of a 1-year-old male dog.     

江苏地区犬细小病毒的分离与变异分析

2007-2013年,在江苏省扬州市和苏州市的病犬临床样本中共分离鉴定了11株犬细小病毒(CPV)。经过VP2特定位置氨基酸的比较分析,证明其中9株为CPV-2a,1株为CPV-2b,1株为CPV-2。部分CPV-2a/b分离毒株在VP2蛋白氨基酸位点300位或440位有新的突变,VP2基因进化树分析表明,大部分江苏毒株在同1个独立分支中。     

Estimated frequency of the canine hyperuricosuria mutation in different dog breeds

Background: Hyperuricosuria is a condition that predisposes dogs to urate urolithiasis. A mutation that causes canine hyperuricosuria was previously identified in 3 unrelated dog breeds. The occurrence of the mutation in additional breeds was not determined. Hypothesis/Objectives: Identify additional breeds that have the hyperuricosuria mutation and estimate the mutant allele frequency in those breeds. Animals: Three thousand five hundred and thirty dogs from 127 different breeds were screened for the hyperuricosuria mutation. Methods: DNA samples were genotyped by pyrosequencing and allele‐specific polymerase chain reaction methods. Results: Mutant allele frequencies that range from 0.001 to 0.15 were identified in the American Staffordshire Terrier, Australian Shepherd, German Shepherd Dog, Giant Schnauzer, Parson (Jack) Russell Terrier, Labrador Retriever, Large Munsterlander, Pomeranian, South African Boerboel, and Weimaraner breeds. Conclusions and Clinical Importance: The hyperuricosuria mutation has been identified in several unrelated dog breeds. The mutant allele frequencies vary among breeds and can be used to determine an appropriate breeding plan for each breed. A DNA test is available and may be used by breeders to decrease the mutant allele frequency in breeds that carry the mutation. In addition, veterinarians may use the test as a diagnostic tool to identify the cause of urate urolithiasis.     

Evaluation of an intron deletion in the c-kit gene of canine mast cell tumors

OBJECTIVE: To evaluate molecular abnormalities in the c-kit gene of canine mast cell tumors (MCT) with different grades of cellular differentiation. SAMPLE POPULATION: 31 normal tissue specimens from dogs and 45 canine MCT classified according to grade of cell differentiation. PROCEDURES: Genomic DNA extractions were made from canine MCT and normal tissues. Parts of exon 11, intron 11, and exon 12 of the c-kit gene were amplified by use of polymerase chain reaction. These regions were cloned, sequenced, and compared with GenBank sequences of the National Center for Biotechnology International. A statistical analysis was used to compare sequences from canine MCT and normal tissues. RESULTS: A significantly higher percentage of homozygous intron 11 deletion was found in canine MCT (49%) than in normal tissues (13%). This percentage was also higher in moderately and poorly differentiated MCT, compared with well-differentiated MCT Although no mutations were detected in any of the specimens, a polymorphism at amino acid position 606 of the canine c-kit sequence was found in all the studied sequences. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated a relationship between intron 11 deletion and MCT and the grade of MCT differentiation. We suggest that intron 11 deletion may be implicated in the pathogenesis of MCT and could be used as a marker for diagnosis and prognosis of canine MCT.     

Expression of vascular endothelial growth factor receptor-1 and -2 in normal and diseased canine eyes

Objective To immunohistochemically evaluate expression of vascular endothelial growth factor receptor‐1 (VEGFR1) and ‐2 (VEGFR2) in ocular tissue of healthy dogs and dogs affected with primary glaucoma, uveitic glaucoma, and intraocular neoplasia. Sample population Enucleated globes from five dogs with primary glaucoma, five dogs with uveitic glaucoma, six dogs with intraocular neoplasms and three ophthalmically normal control dogs. Procedure Ocular tissues were obtained from enucleated globes of clinical cases or immediately following euthanasia for control dogs. Tissue sections were stained immunohistochemically for VEGFR1 and VEGFR2 via standard techniques and vascular tissue was qualitatively evaluated. Vascular endothelial VEGFR1 and VEGFR2 expression patterns are reported for normal and diseased ocular tissues. In addition, VEGFR1 and VEGFR2 expression patterns are reported for all normal ocular tissues. Results A constitutive expression pattern was detected for VEGFR1 by ocular vascular endothelial cells as well as nonvascular cells in the cornea, uvea, lens, and retina. VEGFR2 demonstrated limited expression in normal ocular tissue, but was widely expressed in vascular endothelium of diseased eyes, particularly in pre‐iridal fibrovascular membranes. Conclusions The results of this study suggest a role for VEGF receptors in both physiologic and pathologic angiogenesis in canine ocular tissue. Manipulation of this pathway may be a rational consideration for therapeutic intervention in canine ocular disease exhibiting pathologic neovascularization.