禽大肠杆菌外膜蛋白基因C(ompC)的序列分析

根据 Gen Bank中人源大肠杆菌 K- 12外膜蛋白基因 C(omp C)的核苷酸序列设计引物 ,应用 PCR方法从禽大肠杆菌 O2 、O78株及它们的融合双价弱毒菌株 O2 ,78(Norr Chlr)中分别扩增得到 omp C基因 ,序列测定及分析比较发现 ,3个菌株的 om p C基因均由 170 2 nt组成 ,核苷酸序列完全相同 ,只有 1个大的开放性阅读框 (ORF) ,长 10 92 bp,编码由 36 3个氨基酸组成的前 Om p C蛋白 ,前 2 1个氨基酸残基组成信号肽 ,成熟的 Omp C蛋白由 342个氨基酸残基组成 ,Mr为 4 0 0 0 0。其氨基酸序列也完全相同。从基因水平上证明了禽大肠杆菌 O2 、O78株及融合双价弱毒菌株 O2 ,78(NorrChlr)存在相同的外膜蛋白 C抗原 ,从而为进一步研究 Omp C蛋白的免疫原性奠定了基础     

人胰岛素原基因转染绵羊成纤维细胞及细胞株的建立

从动物耳皮肤组织采样 ,采用将组织块剪碎后直接贴附于培养瓶底部的方法进行原代培养 ,该方法使原代细胞出现率及可传代率均达到 10 0 %。根据上皮样细胞和成纤维样细胞贴壁紧实程度的不同 ,用 0 .0 5 %的胰蛋白酶-EDTA对其进行消化 ,可将两种不同类型的细胞进行分离和纯化。通过脂质体介导 ,以BLG -hINS(含乳球蛋白调控基因的人胰岛素原基因 )基因作为目的基因、GFP(绿色荧光蛋白 )基因作为标记基因共转染绵羊成纤维细胞 ,经G - 4 18筛选后 ,得到转染细胞。对转染的细胞分别用单细胞显微操作法和有限稀释法进行细胞克隆 ,两种方法均可得到克隆细胞。选形态正常、生长均匀的 5个细胞克隆进行PCR检测 ,结果 5个克隆均转有GFP基因 ,其中两个转有BLG -hINS基因。高代培养细胞、转染细胞和克隆细胞经核型分析后 ,染色体数目均为 2 7对 ,表明绵羊耳的成纤维细胞建立细胞株后 ,可以作为外源基因转染的有效供体细胞。     

Myofibrillar proteolysis in chick muscle cell cultures during heat stress

The effect of heat stress on protein oxidation and myofibrillar proteolysis in chick myotubes was investigated. Myotubes were incubated at 37 or 41°C for 6 and 24 h. Protein carbonyl content, as an index of protein oxidation, increased more at 41°C than at 37°C for 6 and 24 h incubations. N^τ‐methylhistidine release as an index of myofibrillar proteolysis also increased more at 41°C than at 37°C for 6 and 24 h incubations. Proteasome activity also increased more under those same conditions. Calpain and cathepsin D but not B + L activities showed a greater increase at 41°C than at 37°C for 24 but not the 6 h incubation. These results indicate that heat stress increases protein oxidation and proteasome activity, resulting in an increase in myofibrillar proteolysis for short‐term incubation and, for long‐term incubation, it increases calpain, proteasome and cathepsin D activities, finally accelerating myofibrillar proteolysis in chick myotubes.     

Results from the treatment of advanced stage squamous cell carcinoma of the nasal planum in cats, using a combination of intralesional carboplatin and superficial radiotherapy: a pilot study

Six cats with an advanced stage squamous cell carcinoma (SCC) of the nasal planum were treated with a combination of superficial radiotherapy and intralesional carboplatin therapy. This multimodality protocol was well tolerated by the majority of cats and resulted in complete responses in all cats (100%). Median follow‐up for all cats is 268 days, and the median time‐to‐recurrence, time‐to‐progression and overall survival have not yet been reached. Our study, although limited in number of animals and with a relatively short median follow‐up compared to other studies for this disease, suggests that a combination of radiotherapy and intralesional carboplatin is a useful treatment option for an advanced stage SCC of the nasal planum in cats and warrants further application of the multimodality approach presented here.     

C-reactive protein concentrations in canine acute pancreatitis

Objective: To determine if C‐reactive protein (CRP) concentration is elevated in spontaneously occurring canine acute pancreatitis (AP), and to measure changes in CRP during the course of hospitalization. Design: Prospective study. Setting: Tufts University School of Veterinary Medicine Foster Hospital for Small Animals. Animals: Sixteen client‐owned dogs with AP and 16 healthy controls. Interventions: Blood samples were obtained from the AP group on the day of diagnosis (Day 1), and on Days 3 and 5, unless the dog died or was discharged from the hospital. Blood was obtained from the control dogs once. Measurements and main results: Serum CRP was measured using a commercial immunoassay for each dog with AP and for healthy controls. Day 1 CRP concentrations were significantly higher in the AP group (56.1±12.7 μg/mL) compared with controls (2.8±1.3 μg/mL; P<0.001). For the 7 dogs that had samples collected on all 3 days, the mean CRP concentrations decreased significantly (P=0.043) over the 5 days of measurement. Of the 16 dogs with AP, 14 were discharged from the hospital and 2 were euthanized. Conclusions: Serum CRP concentrations were elevated in this group of 16 dogs with spontaneously occurring AP. In the 7 dogs that had measurements on all 3 days, the mean CRP concentration decreased from the day of diagnosis to the measurement made 5 days later.     

Expression of the tobacco β-1,3-glucanase gene, PR-2d, following induction of SAR with Peronospora tabacina

Systemic acquired resistance (SAR) is induced following inoculation of Peronospora tabacina sporangia into the stems of Nicotiana tabacum plants highly susceptible to the pathogen. Previous results have shown that accumulation of acidic β-1,3-glucanases (PR-2's) following induction of SAR by P. tabacina may contribute to resistance to P. tabacina. We showed that up-regulation of the PR-2 gene, PR-2d, following stem inoculation with P. tabacina, is associated with SAR. Studies using plants transformed with GUS constructs containing the full length promoter from PR-2d or promoter deletions, provided evidence that a previously characterized regulatory element that is involved in response to salicylic acid (SA), may be involved in regulation of PR-2d following induction of SAR with P. tabacina. This work provides evidence that regulation of PR-2 genes during P. tabacina-induced SAR may be similar to regulation of these genes during infection of N-gene tobacco by TMV or following exogenous application of SA, and provides further support for the role of SA in regulation of genes during P. tabacina-induced SAR.     

Isolation of two strains of a new Tombusvirus (Havel river virus, HaRV) from surface waters in Germany

Two virus isolates from water samples — one from a small stream in South Western Germany and another one from the Havel river in North Eastern Germany c. 500 km away, proved to be strains, named S and H, respectively, of a new Tombusvirus for which the name Havel river virus (HaRV) had been suggested previously in a brief account. Immunoelectron microscopical decoration tests and sequence comparisons of the coat proteins indicated that the two HaRV strains are only distantly related to known Tombusviruses. The closest relationships were found to Cucumber necrosis virus. Nothing is known about their natural hosts. Because the S strain of HaRV was isolated in a woody area from a small stream close to its origin, they may be pathogens of trees or wild plants in such habitats.     

Footsteps from Insect Larvae Damage Leaf Surfaces and Initiate Rapid Responses

Plant resistance to insect herbivory involves gene expression in response to wounding and the detection of insect elicitors in oral secretions (Kessler and Baldwin, 2002, Ann. Rev. Plant/ Biol. 53: 299–328). However, crawling insect larvae stimulate the synthesis of 4-aminobutyrate within minutes and imprints of larval footsteps can be visualized within seconds through superoxide production or transient increases in chlorophyll fluorescence (Bown et al., 2002, Plant Physiol. 129: 1430–1434). Here cryo-scanning electron microscopy was used to demonstrate that larval feet, which are equipped with a perimeter row of hook-like crochets, damage leaf tissue and result in larval footprints. Staining for cell death shows that areas of wounding correspond to footsteps detected through increased chlorophyll fluorescence. Superoxide production in response to footsteps was inhibited by diphenyleneiodonium, an inhibitor of the plasma membrane NADPH oxidase enzyme. Inhibition of superoxide production, however, did not eliminate the detection of cell death. The results demonstrate that larval footsteps damage leaf tissue, and initiate rapid local responses which are not dependent on herbivory or oral secretions. It is proposed that superoxide production at the wound site prevents opportunistic pathogen infection.     

山羊生精上皮细胞分离研究

采用四种酶法分离二月龄关中奶山羊生精上皮细胞,A组:胶原酶Ⅳ;B组:胶原酶Ⅳ +透明质酸酶;C组:胶原酶Ⅳ+透明质酸酶+胰蛋白酶与EDTA;D组:胶原酶Ⅳ+透明质酸酶+胰蛋白酶与EDTA+DnaseI。结果二月龄羔羊曲细精管主要包含Sertoli细胞、精原细胞和管周排列的肌样细胞。每 200mg睾丸实质收获生精上皮细胞总数 (×105 )A,B,C,D组分别为 6. 91±0. 68, 6. 67±0. 80, 5. 94±0. 81, 5. 74±0. 82;细胞存活率 (% )分别为 76. 82±3. 80, 75. 96±1. 61, 94. 19±2. 89, 92. 32±1. 97;圆形细胞比率 (% )分别为 17. 54±1. 68, 16. 70±1. 46, 23. 64±1. 72, 22. 90±2. 38;细胞贴壁率(% )分别为 5. 93±0. 36, 5. 81±0. 54, 26. 94±1. 54, 25. 00±1. 74。C,D组不但组织块解离效果及细胞离散程度均较A,B好,且原代培养细胞团块数少。多重比较C,D组显著优于A,B组 (P<0.05);C组与D组差异不显著(P>0.05),但与A,B组比较细胞存活率、圆形细胞比率及贴壁率差异极显著(P<0.01)。结论认为C组是较简单有效的山羊生精上皮细胞分离酶组合。