海雀稗SRAP-PCR分子标记体系优化及引物筛选

以4个代表性海雀稗(Paspalum vaginatum)种质的叶片DNA为模板,采用L9(34)正交试验设计,对影响海雀稗SRAP-PCR反应的Mg~(2+)、dNTP、引物和TaqDNA聚合酶的用量进行了优化,并比较了不同浓度模板DNA对扩增的影响,以确定适合海雀稗的SRAP-PCR最佳反应体系。结果表明:海雀稗SRAP-PCR最佳反应体系为10×PCR buffer 1μL、模板DNA50ng、Mg2+2.5mmol·L~(-1)、dNTP150μmol·L~(-1)、引物0.4μmol·L~(-1)、TaqDNA聚合酶1.0U,总体积10μL。利用该反应体系从100对引物中筛选出可扩增清晰条带的引物83对,在83对引物中选出多态性丰富的引物44对。SRAP-PCR反应体系的优化及引物筛选,为今后利用SRAP标记技术进行海雀稗遗传多样性研究、基因定位、遗传图谱的构建以及分子标记辅助育种提供技术支持。     

正交设计优化假俭草SRAP-PCR反应体系及引物筛选

以假俭草叶片DNA为模板,采用正交试验设计,以Mg2 、dNTP、引物和Taq DNA聚合酶4种因素3个水平,对假俭草SRAP反应体系进行研究,并比较了不同浓度模板DNA对扩增效果的影响,建立了假俭草的SRAP最佳反应体系.结果表明,假俭草SRAP-PCR最佳反应体系为:2 μL 10譖CR buffer、60 ng模板DNA、Mg2 1.50 mmol/L、dNTP 260 μmol/L、引物0.25 μmol/L、Taq DNA聚合酶0.5 U,总体积为20 μL.各因素对扩增反应结果均有不同影响,其中以Mg2 浓度影响最大,Taq DNA聚合酶的影响最小.运用该体系对4份假俭草种源进行验证,证明该体系稳定可靠,并从45个SRAP引物组合中筛选出扩增条带清晰、多态性丰富的26个引物组合.这一体系的建立及多态性引物组合的筛选为今后利用SRAP标记技术进行假俭草分子遗传学研究提供了科学依据.     

利用正交设计优化羊食道口线虫SRAP-PCR反应体系

本研究利用正交设计L16(4)对羊食道口线虫SRAP-PCR反应体系的4因素:Taq酶、Mg2+、dNTPs、引物进行摸索。结果表明:各因素的不同水平对SRAP-PCR反应结果都有显著的影响,正交设计组合4中扩增的条带最清晰;羊食道口线虫SRAP-PCR最佳反应条件:最佳引物给合为Me1/Em7,最佳反应体系为25μL:2.5μL 10×PCR buffer、20 ng模板DNA、Mg2+(25 mmol/L)3μL、dNTPs(2.5 mmol/L/μL)3μL、引物(10 pmol/μL)4μL、rTaq DNA聚合酶(5U/μL)0.2μL、灭菌ddH20补足。本研究结果为进一步采用SRAP技术来研究食道口线虫遗传进化奠定了坚实基础。     

杂花苜蓿SRAP-PCR反应体系的正交优化

利用正交设计L16(45)对杂花苜蓿(Medicago varia Martin.)SRAP-PCR反应体系的5因素(Taq酶、Mg2+、模板DNA、dNTP、引物)在4个水平上进行优化试验,旨在建立一套适用于苜蓿属(Medicago L.)种质资源的SRAP标记技术体系.结果表明:各因素对PCR反应结果都有显著影响,由F值可知,dNTP的量对反应结果影响最大,其次是Taq酶的量,模版DNA浓度的影响最小,各因素水平的变化对PCR反应的影响依次为:dNTP>Taq酶>Mg2+>引物>模板DNA;筛选出各反应因素的最佳水平,建立杂花苜蓿SRAP-PCR反应的最佳体系(25 μL )为:dNTP 2 μL( 0.20 mmol·L-1),Taq DNA 聚合酶0.3 μL (1.50 U), Mg2+3 μL(1.50 mmol·L-1),上、下游引物各1 μL (0.40 μmol·L-1),50 ng模板DNA1 μL,10×PCR buffer 2.5 μL (不含Mg2+).这一优化体系的建立,为今后利用SRAP标记技术进行苜蓿属植物遗传图谱构建、遗传多样性分析及种质资源鉴定等研究奠定了技术基础.     

野火球RAPD反应体系优化研究初报

为建立一个稳定的野火球RAPD反应体系,对RAPD反应条件中模板DNA用量、Taq DNA聚合酶用量、随机引物浓度、Mg2 浓度和dNTP浓度等各项参数进行筛选比较,创建其最佳反应体系为;250μl反应体系中,模板DNA51ng,Taq DNA聚合酶0.5U.引物浓度0.32μmol/L,Mg2 浓度1.2mmol/L,dNTP浓度0.20mmol/L,10×PCR Buffer 1.0μl,ddH2O19μl.     

正交设计优化草地早熟禾SRAP-PCR反应体系及引物筛选

采用L9(34)正交试验设计方法,对草地早熟禾(Poa pratensis)基因组DNA SRAP PCR反应体系中的Taq DNA聚合酶、Mg2+、引物及dNTP四因素的用量进行优化,并比较不同模板DNA用量对扩增的影响,建立草地早熟禾SRAP PCR最佳反应体系,同时,利用该体系对SRAP引物进行筛选。结果表明,草地早熟禾SRAP PCR最佳反应体系为Taq DNA聚合酶1.0 U、Mg2+ 1.75 mmol·L-1、引物0.25 μmol·L-1、dNTP 220 μmol·L-1、40 ng模板DNA、2 μL 10×PCR buffer,总体积20 μL。运用该体系从100对SRAP引物中筛选出43对引物能够产生清晰稳定的扩增条带且多态性丰富。优化体系的建立及引物的筛选可为今后利用SRAP标记技术对草地早熟禾进行遗传多样性分析、图谱构建、种质资源鉴定奠定技术基础。     

杨桃SCoT标记PCR反应体系建立与验证

用单因素设计法对影响杨桃SCoT-PCR反应体系的主要因素Mg2+, dNTPs,引物, Taq DNA 聚合酶及DNA 模板浓度进行优化。结果表明，适于杨桃研究的SCoT-PCR最佳反应体系为：总体积为20μL的反应体系中，含2.5mmol/L Mg2+，0.3mmol/L dNTPs，30mg/L模板DNA，1.00μmol/L引物和0.4U Taq DNA聚合酶。用不同引物及杨桃DNA对该体系进行验证，扩增条带清晰，结果稳定可靠，证明该反应体系适用于杨桃SCoT-PCR扩增。     

克氏针茅ISSR-PCR反应体系的建立与优化

采用正交试验与单因素设计相结合的方法,对克氏针茅ISSR-PCR反应体系中的5种主要因素(模板DNA、Mg2 、TaqDNA聚合酶、dNTP及引物)进行优化筛选,确立了适合克氏针茅ISSR分析的优化反应体系。在25μL反应体系中各反应成分为:模板DNA 50 ng,Mg2 1.5 mmol/L,dNTP 0.2 mmol/L,引物0.8μmol/L,TaqDNA聚合酶1 U,10×buffer 2.5μL。这一体系的建立为今后利用ISSR技术进行克氏针茅遗传多样性分析、遗传图谱构建和基因定位奠定了技术基础。     

燕麦ISSR反应体系的建立与优化

以10个燕麦品种的基因组DNA为模板,从退火温度、TaqDNA聚合酶、Mg2+、dNTP、引物5个方面对ISSR分子标记体系进行摸索并设计优化试验,建立了一套燕麦ISSR优化反应体系,即:25μl反应体系中,18.3μlddH2O,2.5μl 10×buffer,2.0μmol/L Mg2+,250μmol/L dNTP,1.5 U Taq DNA聚合酶,0.3μmol/L引物,10ng DNA模板。反应程序为:94℃预变性3 min;94℃变性30s,55℃退火30s,72℃延伸1 min,34个循环;72℃延伸5 min,4℃保存。     

正交设计优化崇明白山羊SSR-PCR反应体系

以崇明白山羊基因组DNA为模板,利用正交试验设计L16(45),对影响崇明白山羊SSR-PCR反应体系的5个因素(Taq酶、Mg2+、模板DNA、dNTP及引物的浓度)进行优化。结合单因素完全随机试验筛选各因素的最佳水平,针对MCB3224引物建立了崇明白山羊SSR-PCR的最佳反应体系。结果表明:在20μL的反应体系中,5个因素的最佳水平分别为Taq DNA聚合酶0.5 U、Mg2+2.5 mmol/L、模板DNA 30 ng、dNTP 0.20 mmol/L,引物0.8 mol/L;引物的最佳退火温度为57.3℃。利用16份崇明白山羊的DNA模板和2个微卫星引物验证此反应体系,聚丙烯酰胺凝胶电泳,结果显示该体系稳定可靠且普适性较好。     

Feline intracranial neoplasia: retrospective review of 160 cases (1985-2001)

The purpose of this study was to determine the frequency of different tumor types within a large cohort of cats with intracranial neoplasia and to attempt to correlate signalment, tumor size and location, and survival time for each tumor. Medical records of 160 cats with confirmed intracranial neoplasia evaluated between 1985 and 2001 were reviewed. Parameters evaluated included age, sex, breed, FeLV/FIV status, clinical signs, duration of signs, number of tumors, tumor location(s), imaging results, treatment, survival times, and histopathologic diagnosis. Most of the cats were older (11.3 +/- 3.8 years). Primary tumors accounted for 70.6% of cases. Metastasis and direct extension of secondary tumors accounted for only 5.6 and 3.8% of cases, respectively. Twelve cats (7.5%) had 2 or more discrete tumors of the same type, whereas 16 cats (10.0%) had 2 different types of intracranial tumors. The most common tumor types were meningioma (n = 93, 58.1%), lymphoma (n = 23, 14.4%), pituitary tumors (n = 14, 8.8%), and gliomas (n = 12, 7.5%). The most common neurological signs were altered consciousness (n = 42, 26.2%), circling (n = 36, 22.5%), and seizures (n = 36, 22.5%). Cats without specific neurological signs were common (n = 34, 21.2%). The tumor was considered an incidental finding in 30 (18.8%) cats. In addition to expected relationships (eg, meninges and meningioma, pituitary and pituitary tumors), we found that lesion location was predictive of tumor type with diffuse cerebral or brainstem involvement predictive of lymphoma and third ventricle involvement predictive of meningioma.     

The Prevalence and Intensity of Helminth and Coccidial Infections in Dairy Cattle in Central Kenya

A survey of gastrointestinal parasite infections of young (<6 months old), immature (6–12 months old) and adult (>12 months old) dairy cattle on 16 farms in Kiambu District, Kenya was conducted during a dry season (September 1991 to January 1992) and during a wet season (March to July 1992). The survey was based on monthly coproparasitological examination of cohorts and worm counts in tracer calves. The effects of age, sex, farm and season on the prevalence and intensity of helminth and coccidial infections were determined. Faecal egg and oocyst counts revealed that the overall prevalences were: strongyles (including trichostrongyles) (85.5%), liver flukes (Fasciola gigantica) (34.0%), coccidia (30.9%) and tapeworms (9.6%). Eight species of the protozoan Eimeria were identified, the most prevalent species being E. bovis and E. zuernii. The most prevalent nematode genera were Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus. Season, farm and age of the animals had a significant (p<0.05) influence on the intensity of infection with strongyles, liver flukes and coccidia, whereas the sex of the animals had no significant (p>0.05) effect on the prevalence or intensity of infections. A higher intensity of infection with strongyles and coccidia was found in the wet season than in the dry season (p<0.05). The age-specific intensity was in the following order: for strongyles, immature animals of 6–12 months of age had the highest egg counts, followed by young calves and adults. Calves had significantly (p<0.05) higher oocyst counts than immatures or adults. Liver fluke egg counts did not differ significant (p>0.05) between immatures and adult cattle.     

Hypothalamic dopamine is required for salsolinol‐induced prolactin secretion in goats

The aim of the present study was to clarify the relationship between hypothalamic dopamine (DA) and salsolinol (SAL) for the secretion of prolactin (PRL) in goats. SAL or thyrotropin‐releasing hormone (TRH) was intravenously injected into female goats treated with or without the D₂ DA receptor antagonist haloperidol (Hal), which crosses the blood‐brain barrier, and the PRL‐releasing response to SAL was compared with that to TRH. PRL‐releasing responses to SAL, Hal, and Hal plus SAL were also examined after a pretreatment to augment central DA using carbidopa (Carbi) and L‐dopa. The PRL‐releasing response to Hal alone was greater than that to SAL or TRH alone. The PRL‐releasing response to Hal plus SAL was similar to that of Hal alone. In contrast, the PRL‐releasing response to Hal plus TRH was greater than that to TRH or Hal alone. The treatment with Carbi plus L‐dopa inhibited SAL‐ and Hal‐induced PRL secretion. The inhibition of the PRL‐releasing response to SAL disappeared when SAL was injected with Hal. These results indicate that the mechanisms underlying the SAL‐induced PRL response differ from those of TRH, and suggest that hypothalamic DA and its synthesis is associated in part with SAL‐induced PRL secretion in goats.     

Laboratory diagnosis of respiratory diseases of cattle

The authors describe the procedure of laboratory diagnosis for bovine respiratory diseases: direct diagnosis by isolation and for identification of bacteria or viruses and indirect diagnosis by serological methods. They specify the restraints and limits of this diagnosis and the significance results which are obtained and connected with knowledge of anamnestic information.     

The effect of peri-parturient anthelmintic treatment on the productivity of dairy cattle in subtropical western India

Trials with fenbendazole (Panacur, Hoechst India Ltd) were carried out on two commercial farms in subtropical western India to study the response and economics of nematode treatment in adult dairy cows. Milk yield, lactation length, time to first oestrus and worm egg output were monitored in treated and control groups. Treatment reduced the egg count considerably. Treated cows produced 142 litres more milk over 100 days (p<0.05), with extension of lactation length and advancement of time to first oestrus. The economic gain in terms of milk yield far outweighed the cost of anthelmintic used.     

An Evaluation of the Acceptability as Forage of Some Nutritive and Antinutritive Components and of the Dry Matter Degradation Profiles of Five Species of Ficus

The suitability of five species of Ficus, F. mucoso, F. thonningii, F. polita, F. religiosa and F. benjamina, for feeding ruminant livestock was studied. The nutritive and antinutritive components were determined and the acceptability of the forages to livestock was assessed using a cafeteria method in 8 adult small ruminants. Also, the degradation potential of the dry matter of the five Ficus species was investigated using 3 fistulated sheep. The crude protein (CP) content of the Ficus species varied significantly (p < 0.05), ranging from 130 to 180 g/kg DM. The content of neutral detergent fibre (NDF) also varied significantly (p < 0.05), ranging from 650 to 710 g/kg DM. The organic matter (OM) contents did not differ significantly (p > 0.05), having a mean value of 916 g/kg DM. The concentrations of tannin, saponin, phytic acid and oxalic acid were low. The acceptability of the forage was similar for four of the species but poor for F. polita. The extent of DM degradation varied significant among the Ficus species, the ranking order being F. benjamina > F. thonningii > F. mucoso > F. religiosa > F. polita.