Molecular docking of alkaloid compounds with the matrix metalloproteinase 2

Matrix metalloproteinase protein-2 (MMP-2) is linked to the human oral squamous cell carcinoma. Therefore, it is of interest to design new inhibitors for MMP-2 to combat the disease. Thus, we document the molecular docking features of Aristolochic acid, Cryptopleurine, Epipodophyllotoxin, and Fagaronine with MMP-2 for further consideration.     

Field trials and tribulations--making sense of the regulations for experimental field trials of transgenic crops in Europe

Transgenic plants that are being developed for commercial cultivation must be tested under field conditions to monitor their effects on surrounding wildlife and conventional crops. Developers also use this opportunity to evaluate the performance of transgenic crops in a typical environment, although this is a matter of commercial necessity rather than regulatory compliance. Most countries have adapted existing regulations or developed new ones to deal specifically with transgenic crops and their commodities. The European Union (EU) is renowned, or perhaps notorious, for having the broadest and most stringent regulations governing such field trials in the world. This reflects its nominal adherence to the precautionary approach, which assumes all transgenic crops carry an inherent risk. Therefore, field trials in the EU need to demonstrate that the risk associated with deploying a transgenic crop has been reduced to the level where it is regarded as acceptable within the narrowly defined limits of the regulations developed and enforced (albeit inconsistently) by national and regional governments, that is, that there is no greater risk than growing an equivalent conventional crop. The involvement of national and regional competent authorities in the decision-making process can add multiple layers of bureaucracy to an already-intricate process. In this review, we use country-based case studies to show how the EU, national and regional regulations are implemented, and we propose strategies that could increase the efficiency of regulation without burdening developers with further unnecessary bureaucracy.     

Failure to detect interfering agents in febrile respiratory illnesses

Throat swab specimens from 57 military recruits with febrile respiratory illness in whom no virologic or serologic evidence of infection with respiratory viruses orMycoplasma pneumoniae was found were inoculated and passaged into fresh cultures of monkey kidney cells and continuous lines of human, simian and rabbit cells. Cultures of second passage were challenged with poliovirus type 2 or vaccinia virus and subsequently tested for interference by reading of cytopathic effect and titration of challenge virus. No interference was shown.     

Microtubule configurations and nuclear DNA synthesis during initiation of suspensor-bearing embryos from Brassica napus cv. Topas microspores

In the new Brassica napus microspore culture system, wherein embryos with suspensors are formed, ab initio mimics zygotic embryogenesis. The system provides a powerful in vitro tool for studying the diverse developmental processes that take place during early stages of plant embryogenesis. Here, we studied in this new culture system both the temporal and spatial distribution of nuclear DNA synthesis places and the organization of the microtubular (MT) cytoskeleton, which were visualized with a refined whole mount immunolocalization technology and 3D confocal laser scanning microscopy. A ‘mild’ heat stress induced microspores to elongate, to rearrange their MT cytoskeleton and to re-enter the cell cycle and perform a predictable sequence of divisions. These events led to the formation of a filamentous suspensor-like structure, of which the distal tip cell gave rise to the embryo proper. Cells of the developing pro-embryo characterized endoplasmic (EMTs) and cortical microtubules (CMTs) in various configurations in the successive stages of the cell cycle. However, the most prominent changes in MT configurations and nuclear DNA replication concerned the first sporophytic division occurring within microspores and the apical cell of the pro-embryo. Microspore embryogenesis was preceded by pre-prophase band formation and DNA synthesis. The apical cell of the pro-embryo exhibited a random organization of CMTs and, in relation to this, isotropic expansion occurred, mimicking the development of the apical cell of the zygotic situation. Moreover, the apical cell entered the S phase shortly before it divided transversally at the stage that the suspensor was 3–8 celled.     

Solution-based targeted genomic enrichment for precious DNA samples

ABSTRACT: BACKGROUND: Solution-based targeted genomic enrichment (TGE) protocols permit selective sequencing of genomic regions of interest on a massively parallel scale. These protocols could be improved by: 1) modifying or eliminating time consuming steps; 2) increasing yield to reduce input DNA and excessive PCR cycling; and 3) enhancing reproducible. RESULTS: We developed a solution-based TGE method for downstream Illumina sequencing in a non-automated workflow, adding standard Illumina barcode indexes during the post-hybridization amplification to allow for sample pooling prior to sequencing. The method utilizes Agilent SureSelect baits, primers and hybridization reagents for the capture, off-the-shelf reagents for the library preparation steps, and adaptor oligonucleotides for Illumina paired-end sequencing purchased directly from an oligonucleotide manufacturing company. CONCLUSIONS: This solution-based TGE method for Illumina sequencing is optimized for small- or medium-sized laboratories and addresses the weaknesses of standard protocols by reducing the amount of input DNA required, increasing capture yield, optimizing efficiency, and improving reproducibility.     

Effect of neutralizing sera on factor x-mediated adenovirus serotype 5 gene transfer

The deployment of adenovirus serotype 5 (Ad5)-based vectors is hampered by preexisting immunity. When such vectors are delivered intravenously, hepatocyte transduction is mediated by the hexon-coagulation factor X (FX) interaction. Here, we demonstrate that human sera efficiently block FX-mediated cellular binding and transduction of Ad5-based vectors in vitro. Neutralizing activity correlated well with the ability to inhibit Ad5-mediated liver transduction, suggesting that prescreening patient sera in this manner accurately predicts the efficacy of Ad5-based gene therapies. Neutralization in vitro can be partially bypassed by pseudotyping with Ad45 fiber protein, indicating that a proportion of neutralizing antibodies are directed against the Ad5 fiber.     

Successful development of a shed-microspore culture protocol for doubled haploid production in Indonesian hot pepper (Capsicum annuum L.)

Various systems of anther and microspore cultures were studied to establish an efficient doubled haploid production method for Indonesian hot pepper (Capsicum annuum L.). A shed-microspore culture protocol was developed which outperformed all the previously reported methods of haploid production in pepper. The critical factors of the protocol are: selection of flower buds with more than 50% late unicellular microspores, a 1 day 4°C pretreatment of the buds, followed by culture of the anthers in double-layer medium system for 1 week at 9°C and thereafter at 28°C in continuous darkness. The medium contained Nitsch components and 2% maltose, with 1% activated charcoal in the solid under layer and 2.5 μM zeatin and 5 μM indole-3-acetic acid in the liquid upper layer. All the ten genotypes of hot pepper tested, responded to this protocol. The best genotypes produced four to seven plants per original flower bud. This protocol can be used as a potential tool for producing doubled haploid plants for hot pepper breeding.     

Neutralizing antibodies to adenovirus serotype 5 vaccine vectors are directed primarily against the adenovirus hexon protein

The utility of recombinant adenovirus serotype 5 (rAd5) vector-based vaccines for HIV-1 and other pathogens will likely be limited by the high prevalence of pre-existing Ad5-specific neutralizing Abs (NAbs) in human populations. However, the immunodominant targets of Ad5-specific NAbs in humans remain poorly characterized. In this study, we assess the titers and primary determinants of Ad5-specific NAbs in individuals from both the United States and the developing world. Importantly, median Ad5-specific NAb titers were >10-fold higher in sub-Saharan Africa compared with the United States. Moreover, hexon-specific NAb titers were 4- to 10-fold higher than fiber-specific NAb titers in these cohorts by virus neutralization assays using capsid chimeric viruses. We next performed adoptive transfer studies in mice to evaluate the functional capacity of hexon- and fiber-specific NAbs to suppress the immunogenicity of a prototype rAd5-Env vaccine. Hexon-specific NAbs were remarkably efficient at suppressing Env-specific immune responses elicited by the rAd5 vaccine. In contrast, fiber-specific NAbs exerted only minimal suppressive effects on rAd5 vaccine immunogenicity. These data demonstrate that functionally significant Ad5-specific NAbs are directed primarily against the Ad5 hexon protein in both humans and mice. These studies suggest a potential strategy for engineering novel Ad5 vectors to evade dominant Ad5-specific NAbs.