Vectors for multi-color bimolecular fluorescence complementation to investigate protein-protein interactions in living plant cells

<p>Abstract</p> <p>Background</p> <p>The investigation of protein-protein interactions is important for characterizing protein function. Bimolecular fluorescence complementation (BiFC) has recently gained interest as a relatively easy and inexpensive method to visualize protein-protein interactions in living cells. BiFC uses "split YFP" tags on proteins to detect interactions: If the tagged proteins interact, they may bring the two split fluorophore components together such that they can fold and reconstitute fluorescence. The sites of interaction can be monitored using epifluorescence or confocal microscopy. However, "conventional" BiFC can investigate interactions only between two proteins at a time. There are instances when one may wish to offer a particular "bait" protein to several "prey" proteins simultaneously. Preferential interaction of the bait protein with one of the prey proteins, or different sites of interaction between the bait protein and multiple prey proteins, may thus be observed.</p> <p>Results</p> <p>We have constructed a series of gene expression vectors, based upon the pSAT series of vectors, to facilitate the practice of multi-color BiFC. The bait protein is tagged with the C-terminal portion of CFP (cCFP), and prey proteins are tagged with the N-terminal portions of either Venus (nVenus) or Cerulean (nCerulean). Interaction of cCFP-tagged proteins with nVenus-tagged proteins generates yellow fluorescence, whereas interaction of cCFP-tagged proteins with nCerulean-tagged proteins generates blue fluorescence. Additional expression of mCherry indicates transfected cells and sub-cellular structures. Using this system, we have determined in both tobacco BY-2 protoplasts and in onion epidermal cells that <it>Agrobacterium </it>VirE2 protein interacts with the <it>Arabidopsis </it>nuclear transport adapter protein importin α-1 in the cytoplasm, whereas interaction of VirE2 with a different importin α isoform, importin α-4, occurs predominantly in the nucleus.</p> <p>Conclusion</p> <p>Multi-color BiFC is a useful technique to determine interactions simultaneously between a given" bait" protein and multiple "prey" proteins in living plant cells. The vectors we have constructed and tested will facilitate the study of protein-protein interactions in many different plant systems.</p

Effects of immersing treatment of curcumin and piperine combined with vacuum packaging on the quality of salmon (Salmo salar) during cold chain logistics

In order to study the effects of the compound preservatives (curcumin and piperine (CP)) and vacuum packaging (VP) on the quality of salmon during cold chain logistics suffered from temperature abuse, the physiochemical indexes (texture, water holding capacity (WHC), total volatile basic nitrogen (TVB-N), thiobarbituric acid reactive substances (TBARS), free amino acids (FAA) contents), microbial indicators (total mesophilic bacteria count (MBC), total psychrotrophic bacteria count (PBC), H2S-producing bacteria count (HSBC)) were determined, and the moisture changes were explored by near-infrared (NIR) spectroscopy and low-field nuclear magnetic resonance (LF-NMR). The results showed that the treatment of curcumin and piperine in combination with vacuum packaging could maintain the quality of salmon suffered from temperature abuse most effectively. At the end of storage, the MBC of VP+CP was only 4.95 log CFU/g, which was about 1 log CFU/g lower than the control sample stored at the same condition. The combined treatment also retarded the increase of TVB-N, TBARS, and the decrease of hardness, springiness, and a* value, as well as water migration in salmon, contributing to higher water holding capacity and better appearance. Besides, VP+CP retarded the decrease of free glutamate, which contributed to umami taste. Due to the biological activity and safety of the preserves, the combined treatment could be a promising method for preservation of seafood.Peer reviewe

Ammonium 2-(2,4-dichloro­phen­oxy)acetate hemihydrate

The title compound, NH4 +·C8H7Cl2O6 −·0.5H2O, was prepared by the reaction of 2-(2,4-dichloro­phen­oxy)­acetic acid and ammonia in water at 367 K. The mol­ecular structure and packing are stabilized by N—H⋯O and O—H⋯O inter­molecular hydrogen-bond inter­actions

Equine Arteritis Virus Does Not Induce Interferon Production in Equine Endothelial Cells: Identification of Nonstructural Protein 1 as a Main Interferon Antagonist

The objective of this study was to investigate the effect of equine arteritis virus (EAV) on type I interferon (IFN) production. Equine endothelial cells (EECs) were infected with the virulent Bucyrus strain (VBS) of EAV and expression of IFN-β was measured at mRNA and protein levels by quantitative real-time RT-PCR and IFN bioassay using vesicular stomatitis virus expressing the green fluorescence protein (VSV-GFP), respectively. Quantitative RT-PCR results showed that IFN-β mRNA levels in EECs infected with EAV VBS were not increased compared to those in mock-infected cells. Consistent with quantitative RT-PCR, Sendai virus- (SeV-) induced type I IFN production was inhibited by EAV infection. Using an IFN-β promoter-luciferase reporter assay, we subsequently demonstrated that EAV nsps 1, 2, and 11 had the capability to inhibit type I IFN activation. Of these three nsps, nsp1 exhibited the strongest inhibitory effect. Taken together, these data demonstrate that EAV has the ability to suppress the type I IFN production in EECs and nsp1 may play a critical role to subvert the equine innate immune response

Retinal vessel diameter changes induced by transient high perfusion pressure

AIM: To investigate the effects of transient high perfusion pressure on the retinal vessel diameter and retinal ganglion cells. METHODS: The animals were divided into four groups according to different infusion pressure and infusion time (60 mm Hg-3min, 60 mm Hg-5min, 100 mm Hg-3min, 100 mm Hg-5min). Each group consisted of six rabbits. The left eye was used as the experimental eye and the right as a control. Retinal vascular diameters were evaluated before, during infusion, immediately after infusion, 5min, 10min and 30min after infusion based on the fundus photographs. Blood pressure was monitored during infusion. The eyes were removed after 24h. Damage to retinal ganglion cell (RGC) was analyzed by histology. RESULTS:Retina became whiten and papilla optic was pale during perfusion. Measurements showed significant decrease in retinal artery and vein diameter during perfusion in all of the four groups at the proximal of the edge of the optic disc. The changes were significant in the 100 mm Hg-3min group and 100 mm Hg-5min group compared with 60 mm Hg-3min group (P1=0.025, P2=0.000). The diameters in all the groups recovered completely after 30min of reperfusion. The number of RGC showed no significant changes at the IOP in 100 mm Hg with 5min compared with contralateral untreated eye (P＞0.05). CONCLUSION: Transient fluctuations during infusion lead to temporal changes of retinal vessels, which could affect the retinal blood circulation. The RGCs were not affected by this transient fluctuation. Further studies are necessary to evaluate the effect of pressure during real-time phacoemusification on retinal blood circulation

Avian malaria parasite infections do not affect personality in the chestnut thrush (Turdus rubrocanus) on the Qinghai-Tibet Plateau

Personality traits, the consistent individual behavioral differences, are currently gaining much attention in studies of natural bird populations. However, associations between personality traits and parasite infections are not often investigated. Even less attention has been given to studies of birds in the high-elevation region such as the Tibetan plateau. This research aims to examine the relationship between avian malaria parasites and two personality traits in a population of the Chestnut Thrush (Turdus rubrocanus) breed in the Tibetan plateau. Our results revealed no evidence of sex bias in malaria parasite prevalence. Furthermore, we found no effect of infection status on two personality scores: activity and boldness. Additionally, no effects on the activity level or boldness were observed for different parasite lineages of Haemoproteus, Leucocytozoon, the sex of the birds, or their interactions. Similarly, we did not find any relationship between activity level and boldness with nestling numbers, sex, or their interactions. Notably, individuals with a larger number of offspring tended to display greater boldness. Our findings indicate that blood parasite infections are common in this population but do not significantly impact the personality of the birds

Surgical treatment of multivalvular endocarditis: Twenty-one–year single center experience

ObjectiveLittle information is available about surgical outcomes in patients with multivalvular endocarditis. The aim of this article is to review the 21-year experience with surgical treatment of patients with multivalvular endocarditis at our institution and, in particular, to determine the incidence, pathologic status, diagnosis, surgical strategies, and outcomes of patients with this disease.MethodsFrom January 1986 to December 2006, a total of 48 patients (40 men, 8 women), with a mean age of 42 ± 12 years, underwent surgery for multivalvular endocarditis. Endocarditis was active in 32 patients and healed in 16. Preoperative transthoracic echocardiographic evaluation was performed in all 48 patients with addition of transesophageal echocardiography in 22 (45.8%). Intraoperative findings showed that the endocarditis involved mostly the mitral and aortic valves (40/48 patients). Triple or quadruple valve involvement was found in 1 and 2 patients, respectively. Preoperative, perioperative, and postoperative data were retrospectively analyzed and risk factors for early and late survival were determined.ResultsIn only 24 (50.0%) patients was multivalvular endocarditis diagnosed by preoperative transthoracic echocardiography; 17 (77.3%) patients had multivalvular endocarditis confirmed by preoperative transesophageal echocardiography. The 30-day hospital mortality was 12.5% (n = 6). Preoperative renal failure, New York Heart Association class IV, and emergency surgery were identified as independent risk factors for hospital mortality. Overall long-term survival was 74% ± 6% at 5 years and 62% ± 3% at 10 years. Multivariate analysis revealed that renal failure and recurrent endocarditis were associated with increased late mortality. Ten-year freedom from recurrent endocarditis was 74% ± 5% and 10-year freedom from reoperation was 73% ± 6%.ConclusionsIn our institution, multivalvular endocarditis was diagnosed by transthoracic echocardiography in only half of the patients. Intraoperative transesophageal echocardiography provided a more effective means to identify this disease. Radical resection of all infected tissues for patients with multivalvular endocarditis and additional intraoperative interventions, depending on the intraoperative pathologic condition, produced satisfactory in-hospital and long-term results, similar to those in patients with a single infected heart valve

Zhi-Bai-Di-Huang-Wan, a classic Chinese medicinal formula in relieving menopausal symptoms: A multi-centre and controlled trial from UK and China

Background: To explore the effects of Zhi-Bai-Di-Huang-Wan (ZBDHW), a classic formula of Chinese  medicinal herbs in relieving menopausal symptoms in British and Chinese women.Methods and Materials: Between May 2011 and May 2013, 224 Chinese and British women were  divided into a ZBDHW group with 115 cases and a control group with 109 cases. The clinical menopausal  symptoms were assessed by the modified Kupperman Index Scale. The serum levels of follicle stimulating hormone (FSH) and estradiol (E2) were respectively detected before and after the treatment. Results: After 12 weeks of treatment, both groups’ Kupperman index scores markedly decreased  (P&lt;0.05) and no significant difference existed between them (P&gt;0.05). The serum levels of FSH did not change  significantly after the treatment (P&gt;0.05) and no significant difference existed between them (P&gt;0.05). The  serum levels of E2 significantly increased in both of the two groups (P&lt;0.05) and it increased more in the comparison group (P&lt;0.05). No side-effect of the treatment was reported in both of the two groups during the  period of the treatment.Conclusion: The classic Chinese medicinal formula, ZBDHW, showed promise in relieving menopausal  symptoms.Key words: Chinese medicinal herb; Zhi-Bai-Di-Huang-Wan (ZBDHW); menopausal symptom