黄喉拟水龟转铁蛋白基因的克隆以及表达特征分析

转铁蛋白在铁的新陈代谢中起着重要的作用,参与细菌感染后的免疫反应。研究克隆了黄喉拟水龟Tf基因组DNA全长序列,并对Tf基因的序列特征进行了分析。序列分析表明,黄喉拟水龟Tf是由两个相似结构域构成的单一肽链,每个结构域包含两个亚基,它们相互作用形成一个深的、亲水的铁结合位点。其基因组DNA由17个外显子和16个内含子组成,与其他脊椎动物Tf基因结构相似,显示了黄喉拟水龟Tf基因在结构上的保守性。同源性分析表明,黄喉拟水龟的Tf基因与鸟类、爬行类的Tf基因同源性最高,约为75%—97%;与哺乳类Tf基因的同源性约为65%—75%;与爪蟾等两栖类的Tf基因也有一定的同源性。荧光定量PCR结果显示,黄喉拟水龟人工感染粘质沙雷氏菌后,Tf基因在其肝脏、脾脏、肾脏及心脏各组织中的表达量均有上调的趋势,这与其他动物经病原刺激后的表达特征具有相似性。       

中国水龟类的核型与银带带型研究

以血培养细胞为材料,研究了三种水龟的核型与ＮＯＲｓ,结果表明,染色体数均为２ｎ＝５２,核型模式９＋５＋１２,属龟科核型中的原始类型,但眼斑水龟ＮＦ＝７６,而四眼斑水龟和黄喉拟水龟的ＮＦ＝７８。从核型结构看,眼斑水龟与四眼斑水龟相似,而黄喉拟水龟则有较大差别,作者还发现黄喉拟水龟核型的一些重要特征,即其Ａ组Ｎｏ．３和Ｎｏ．６染色体有次缢痕,Ｎ．别,作者还发现黄喉拟水龟核型的一些重要特征,即Ａ组这些是     

黄喉拟水龟摩氏摩根菌的分离鉴定及药敏试验

<正>黄喉拟水龟(Mauremys mutica),俗称石龟、石金钱龟、黄板龟等,为龟科拟水龟属,主要分布于中国江苏、安徽、福建以及越南、日本等。黄喉拟水龟具有食用、药用和观赏价值,近年来野生资源日趋减少,人工养殖发展迅速。细菌性病原感染是黄喉拟水龟养殖过程中常见疾病,目前已报道黏质沙雷氏菌(Serratia marcescens)[1]、松鼠葡萄球菌(Staphylococcus sciuri)[2]、摩氏摩根菌(Morganella morganii)[3]、嗜麦芽窄食单胞菌(Stenotrophmomas     

黄喉拟水龟和乌龟盾片的变异

龟类盾片的数目、形状和排列方式是鉴别属种的重要特征之一。我们在进行皖南龟类区系调查时,采到了盾片的数目和形状与记载的黄喉拟水龟(Mauremysmutica)和乌龟(Chinemysreevsii)差异较大的标本,经研究认为系个体变异。今简单报道如下:一、黄喉拟水龟Mauremysmutica关于黄喉拟水龟的属名,国内外有二种意见:田婉淑、江耀明(1986)和胡淑琴等(1962)采用Clemmysmutica;赵尔宓(1986)采用Mauremysmutica。McDowell(1964)将黄喉水龟(Clemmysmutica)订正为黄喉拟水龟(Mauremysmutica),并有详细的叙述。我们检查了此龟的骨骼,同意McDowell和赵尔…     

黄喉拟水龟（♀）与三线闭壳龟（♂）杂交后代的形态特征及其与父母本的比较研究

本研究通过黄喉拟水龟Mauremys mutica（♀）与三线闭壳龟Cuora trifasciata（♂）进行杂交,成功获得了杂种龟。这说明黄喉拟水龟和三线闭壳龟是可以进行属间远缘杂交的,但杂交组合的受精率及孵化成功率均低于黄喉拟水龟的同种组合。杂种稚龟与黄喉拟水龟稚龟在背甲纹路、腹甲黑斑、四肢和尾腹面的皮肤颜色、喉盾前端形状及起点位置、喉盾宽/喉盾缝长、喉盾缝长/肱盾缝长、股盾缝长/肛盾缝长存在差异。1龄前,杂种龟生长快于黄喉拟水龟。形态特征上,杂种龟头顶部淡棕黄色,头侧眼后有两条黑色纵纹,颈腹部淡黄色;背甲棕色,腹甲浅黄色,每一盾片中间都有边缘呈放射状的黑斑;四肢、尾腹面及裸露皮肤部分为黄褐色。形态可量数据分析显示杂种龟在形态上接近黄喉拟水龟。建立了三种龟的形态判别公式,判别的准确率为100%（p<0.01）,判别分析中贡献最大的4个变量分别是腹甲后半部长/背甲长、喉盾宽/背甲长、肱盾缝长/背甲长、股盾缝长/背甲长,可见三线闭壳龟、黄喉拟水龟与杂交种腹部的腹甲后半部长、喉盾宽、肱盾缝长、股盾缝长等参数可作为鉴别三者的直接依据。本实验的结果对杂种龟的鉴定、龟类的杂交育种及养殖生产实践都有一定的借鉴作用。     

黄喉拟水龟的繁殖生物学研究

1998年4-11月对黄喉拟水龟的繁殖生物学进行了研究,在广东,黄喉拟水龟的产卵高峰期在5-6月,窝卵量1-7枚,平均2.5枚;卵重平均13.95g;长径平均4.34cm;短径平均2.23cm。卵重与卵长径呈正相关关系,在25-32℃、沙床含水量5-10%孵化条件下,孵化时间平衡73.8d,孵化率84.2%。初生稚龟均重9.75g。背甲长平均3.77cm;背甲宽平均2.91cm。稚龟体重与背甲长、宽呈极显著的正相关关系,稚龟经8-11月3个月生长,平均日增重0.149g。稚龟体重、背甲长、宽与日龄呈极显著的正相关关系。稚龟体重与背甲长宽也呈极显著的正相关关系。     

基于微卫星多重PCR技术的黄喉拟水龟亲子鉴定

利用黄喉拟水龟(Mauremys mutica)微卫星标记,筛选出16对微卫星引物,通过优化各引物比例、荧光接头浓度、退火温度和循环次数,建立了基于2组各含8个微卫星位点多重PCR体系的黄喉拟水龟亲子鉴定技术。应用2组微卫星多重PCR体系,通过ABI3130遗传分析仪以及cervus3.0软件对428只黄喉拟水龟进行了个体基因型检测和遗传多样性分析,结果显示,群体的平均等位基因数为14.190,平均多态信息含量为0.748,平均观测杂合度和期望杂合度分别为0.687、0.771。对89只候选母本及296只子代进行亲子鉴定分析,结果显示:在父本信息未知时,母本鉴定率为87%;母本获得的子代个数范围为1-12,个体间表现出巨大的差异,这为选择育种提供了物质基础。黄喉拟水龟多重PCR亲子鉴定技术的建立为群体遗传多样性分析、家系鉴定管理和选择育种提供了有效的技术手段。       

不同饵料养殖黄喉拟水龟效果的研究

用5种不同的饵料(中华鳖配合饲料、蚯蚓、鲢鱼肉、福寿螺肉和河蚌肉)分别投喂黄喉拟水龟幼龟,结果表明,①投喂中华鳖配合饲料的黄喉拟水龟,其增重速度最快,平均日增重和蛋白质效率都最高,且耗料增重比最低,其后依次是: 蚯蚓＞鲢鱼肉＞福寿螺肉＞河蚌肉.②投喂福寿螺肉的黄喉拟水龟,其饵料成本最低,其后依次是: 中华鳖配合饲料＞蚯蚓＞河蚌肉＞鲢鱼肉.③中华鳖配合饲料是养殖黄喉拟水龟较好的饵料,但福寿螺、蚯蚓来源方便,也不失为养殖黄喉拟水龟的好饵料.     

黄喉拟水龟消化道的组织学观察

观察黄喉拟水龟消化道的组织结构.采用常规石蜡切片和HE染色方法对黄喉拟水龟的消化道进行观察.除了口咽腔以外,消化道的管壁是由粘膜层、粘膜下层、肌肉层和外膜组成;各部分的主要区别在于粘膜层,食道和大肠的是复层柱状上皮,胃和小肠的是单层柱状上皮.黄喉拟水龟的舌桔红色,不能伸缩;食管中无食管腺,扩张性强;胃呈囊状,有大量胃腺,腔面皱襞较多;小肠较长,是消化的主要场所,表面有大量的绒毛,在绒毛中可见肠腺;大肠无绒毛,也存在皱襞.     

The Historical Speciation of Mauremys Sensu Lato: Ancestral Area Reconstruction and Interspecific Gene Flow Level Assessment Provide New Insights

Mauremys sensu lato was divided into Mauremys, Chinemys, Ocadia, and Annamemys based on earlier research on morphology. Phylogenetic research on this group has been controversial because of disagreements regarding taxonomy, and the historical speciation is still poorly understood. In this study, 32 individuals of eight species that are widely distributed in Eurasia were collected. The complete mitochondrial (mt) sequences of 14 individuals of eight species were sequenced. Phylogenetic relationships, interspecific divergence times, and ancestral area reconstructions were explored using mt genome data (10,854 bp). Subsequent interspecific gene flow level assessment was performed using five unlinked polymorphic microsatellite loci. The Bayesian and maximum likelihood analyses revealed a paraphyletic relationship among four old genera (Mauremys, Annamemys, Chinemys, and Ocadia) and suggested the four old genera should be merged into the genus (Mauremys). Ancestral area reconstruction and divergence time estimation suggested Southeast Asia may be the area of origin for the common ancestral species of this genus and genetic drift may have played a decisive role in species divergence due to the isolated event of a glacial age. However, M. japonica may have been speciated due to the creation of the island of Japan. The detection of extensive gene flow suggested no vicariance occurred between Asia and Southeast Asia. Inconsistent results between gene flow assessment and phylogenetic analysis revealed the hybrid origin of M. mutica (Southeast Asian). Here ancestral area reconstruction and interspecific gene flow level assessment were first used to explore species origins and evolution of Mauremys sensu lato, which provided new insights on this genus.     

黑颈乌龟两个养殖种群的遗传多样性分析

开展濒危物种养殖群体的遗传多样性研究,对种质资源的保护与利用具有重要意义.选择线粒体COI基因作为本次研究的分子标记,分别从序列组成、单倍型及遗传距离、种群历史动态等方面对黑颈乌龟(Mauremys nigricans)两个养殖种群进行比较分析.结果显示:两个种群的COI序列全长572 bp,A+T含量为55.4％,G...     

The Effect of the Cyp19a1 Gene Methylation Modification on Temperature-dependent Sex Determination of Reeves' Turtle(Mauremys reevesii)

Temperature-dependent sex determination(TSD) is a type of environmental sex determination in which the sex of the embryos depends on the ambient temperature; however,the molecular mechanisms governing this process remain unknown.Aromatase,encoded by the cyp19a1 gene,which converts androgens into estrogens in animals,was considered to be the key gene for TSD.In this study,the 5'-flanking region of the cyp19a1 gene in Reeves' turtle(Mauremys reevesii) was cloned,and the promoter region was identified using the luciferase reporter assay.Then the eggs of Reeves' turtle were incubated at different temperatures(26°C: male-biased temperature; 29°C: non-sex-biased temperature and 32°C: female-biased temperature).During the thermosensitive period,the adrenal kidney gonad complexes(AKG) were sampled.DNA methylation analysis of the AKG samples showed that the promoter region of the cyp19a1 gene was significantly de-methylated in the female-biased temperature regime(P0.01).Quantitative analysis of the cyp19a1 gene and estrogen by q PCR and Elisa assay showed that the expression level of the cyp19a1 gene and estrogen content were both upregulated significantly at the female-biased temperature(P0.01).These results indicated that the de-methylation response of the cyp19a1 gene to incubation temperature,especially at the female-biased temperature,could lead to temperature-specific expression of aromatase and increased estrogen levels,which may further determine gonadal development in Reeves' turtle.These findings provide insights into the genetic mechanisms underlying TSD.     

Interdigitating cells in the thymus of the turtle Mauremys caspica

Summary Interdigitating cells are non-lymphoid elements in the thymus and peripheral, secondary lymphoid organs of higher vertebrates. Their origin and functional significance are a matter of controversy. In the present investigation we analyze, for the first time, the nature of presumptive interdigitating cells of the thymus of an ectothermic vertebrate, the turtle Mauremys caspica. This model is specially useful because of the seasonal variations that affect the reptilian lymphoid organs. Immature pro-interdigitating cells and phagocytosing mature interdigitating cells are described with special emphasis on their ultrastructural characteristics and possible relationships with monocytes and macrophages.     

Ultrastructure of splenic white pulp of the turtle,Mauremys caspica

Summary The ultrastructure of splenic tissue of non-immunized turtles, Mauremys caspica, shows two areas, namely, the white pulp which is lymphoid in nature, and the red pulp which is formed by cell cords and sinusoids. Between both areas there is always a marginal zone with gaps through which cells leak. In the white pulp, there are two blood vessel types; one with muscled walls, and the other showing thinner walls sheathed by reticular cells. Reticular cells constitute a network where there occur dendritic macrophages, lymphoblasts and small and medium lymphocytes. Mature plasma cells are scarce in the white pulp.     

亚硝酸盐暴露对红耳龟与中华条颈龟幼体的慢性毒性效应

对比研究了亚硝酸盐4个浓度NaNO2(0、100、200、500 mg/L)暴露对红耳龟(平均体重13.89 g±1.67 g,n=224)和中华条颈龟(平均体重13.12 g±1.83 g,n=236)机体慢性毒性效应的影响.结果 表明,红耳龟各暴露组肠中亚硝酸盐含量均低于中华条颈龟,且胁迫20 d时差异极显著(P＜...     

The immune response of thymic cells from the turtle Mauremys caspica

In the present work several known mammalian leukocyte functions such as directed mobility, proliferative response to mitogens, antibody-dependent cellular cytotoxicity (ADCC) and natural killer (NK)-mediated cytotoxicity were studied in female and male Mauremys caspica turtles. Since the reptilian thymus shows seasonal variations in its structure, we have performed all the assays along the seasonal cycle. Our results show that thymic cells from M. caspica are able to migrate through a chemo-attractant gradient, to proliferate in response to the mitogens phytohaemagglutinin (PHA) and concanavalin A (Con A), and to kill tumoral target cells by both ADCC-mediated and NK-mediated cytotoxicity. Those functions were differentially affected by the seasonal cycle; in general, in autumn the functions studied showed the smallest values for both sexes, while in summer the highest values of cytotoxicity and chemotaxis were found in females. The proliferative responses to PHA and Con A were higher for both sexes in spring and for females in winter than in the other seasons. In summary, thymic cells from M. caspica show a wide range of immune functions, and these are modulated heterogeneously by the seasonal cycle in both sexes.     

White pulp compartments in the spleen of the turtle Mauremys caspica

Summary The aim of the present study was to analyze the nature of lymphoid and non-lymphoid cellular components occurring in distinct histological compartments of the splenic white pulp of the turtle, Mauremys caspica, in order to define their possible correlations with those of the spleen of higher vertebrates, principally mammals. The white pulp of M.caspica consisted of 3 clearly distinguishable regions: (1) the periateriolar lymphoid sheath, and (2) the inner and (3) the outer zones of the periellipsoidal lymphoid sheath. Reticular cells intimately associated with reticular fibres constituted an extensive meshwork in the periarteriolar lymphoid sheath which housed principally Ig-negative lyphoid cells, mature and immature plasma cells, and interdigitating cells. A few Ig-positive cells were also present in the peripheral region of the periarteriolar lymphoid sheath. The inner and outer zones of the periellipsoidal lymphoid sheath were separated by a discontinuous layer of reticular cell processes. In the inner zone, surface Ig-positive lymphoid cells predominated as well as dendritic cells, resembling ultrastructurally the mammalian follicular dendritic cells, although no germinal centres were found in the turtle spleen. Macrophages, some cytoplasmic Ig-positive cells, and Ig-negative lymphoid cells appeared in the outer zone of the periellipsoidal lymphoid sheath. These results allow us to speculate on a phylogenetic relationship between the periarteriolar lymphoid sheath and the inner and the outer zones of the periellipsoidal lymphoid sheath of the spleen of M. caspica and the periarteriolar lymphoid tissue, the lymphoid follicles and the marginal zone, respectively, of the mammalian splenic white pulp.