Inheritance and expression of a transgene insert in an aneuploid tobacco line

A T-DNA locus comprising nptII, uidA and nos genes — all under the control of the nos promoter (this locus was designated K because it encodes resistance to Kanamycin) - was found to be inherited erratically in a transgenic tobacco line. This anomalous behavior was partially explained following a karyotype analysis of plants representing several generations: these plants were aneuploids, presumably for the K-containing chromosome. During four generations of sexual propagation, transgenic plants that were either trisomic or tetrasomic for the K-containing chromosome (i.e. 2n=49 or 2n=50, respectively) were obtained. The trisomic plants (2n=48+1) were virtually indistinguishable phenotypically from normal euploids (2n=4x=48), whereas the tetrasomic plants (2n=48+2) were smaller, had somewhat misshapen leaves and exhibited reduced fertility. Although the amount of NPTH protein in different trisomic (K--, KK-, KKK) and tetrasomic (KK--, KKK-) plants was generally consistent with a K dosage effect, the genetic behavior of each trisomic — with respect to segregation of Kan^R and marker gene activity in progeny — was unique and not completely explicable by invoking aneuploidy. Specifically, unexpected gains or losses of K could occur, suggesting the formation of double reductional gametes and/or frequent gene conversion at this locus. The susceptibility of K locus marker genes to trans-inactivation in the trisomic and tetrasomic lines was tested by crossing in partially homologous silencing loci. In all transgenotypes tested, the three K marker genes were sensitive to trans-silencing, which was accompanied by methylation in all copies of the nos promoter. In addition to this directed inactivation/methylation, the K locus could also undergo infrequent, spontaneous partial methylation, which produced stable epialleles. In most plants, however, the multiple copies of the nos promoter at this locus remained unmethylated and active through four generations in all transgenotypes examined. The significance of these results for irregular inheritance patterns, aneuploid syndromes and homology-dependent gene silencing is discussed.     

Immobilization ofColeus blumei cells in a column reactor using a spray feeding system

Summary Coleus blumei cells were immobilized in a column reactor packed withLuffa cylindrica pieces. Medium was fed from the top of the column using a spray system and cells maintained high viability for 52 days. Cell growth was slower but rosmarinic acid production was better compared to immobilized cells in the shake flasks.     

The vascular endothelial growth factor (VEGF) isoforms: differential deposition into the subepithelial extracellular matrix and bioactivity of extracellular matrix-bound VEGF.

Vascular endothelial growth factor (VEGF)mRNA undergoes alternative splicing events that generate four different homodimeric isoforms, VEGF121, VEGF165, VEGF189, or VEGF206. VEGF121 is a nonheparin-binding acidic protein, which is freely diffusible. The longer forms, VEGF189 or VEGF206, are highly basic proteins tightly bound to extracellular heparin-containing proteoglycans. VEGF165 has intermediate properties. To determine the localization of VEGF isoforms, transfected human embryonic kidney CEN4 cells expressing VEGF165, VEGF189, or VEGF206 were stained by immunofluorescence with a specific monoclonal antibody. The staining was found in patches and streaks suggestive of extracellular matrix (ECM). VEGF165 was observed largely in Golgi apparatus-like structures. Immunogold labeling of cells expressing VEGF189 or VEGF206 revealed that the staining was localized to the subepithelial ECM. VEGF associated with the ECM was bioactive, because endothelial cells cultured on ECM derived from cells expressing VEGF189 or VEGF206 were markedly stimulated to proliferate. In addition, ECM-bound VEGF can be released into a soluble and bioactive form by heparin or plasmin. ECM-bound VEGF189 and VEGF206 have molecular masses consistent with the intact polypeptides. The ECM may represent an important source of VEGF and angiogenic potential.     

Integrating sample similarities into latent class analysis: a tree-structured shrinkage approach

This paper is concerned with using multivariate binary observations to estimate the probabilities of unobserved classes with scientific meanings. We focus on the setting where additional information about sample similarities is available and represented by a rooted weighted tree. Every leaf in the given tree contains multiple samples. Shorter distances over the tree between the leaves indicate a priori higher similarity in class probability vectors. We propose a novel data integrative extension to classical latent class models with tree-structured shrinkage. The proposed approach enables (1) borrowing of information across leaves, (2) estimating data-driven leaf groups with distinct vectors of class probabilities, and (3) individual-level probabilistic class assignment given the observed multivariate binary measurements. We derive and implement a scalable posterior inference algorithm in a variational Bayes framework. Extensive simulations show more accurate estimation of class probabilities than alternatives that suboptimally use the additional sample similarity information. A zoonotic infectious disease application is used to illustrate the proposed approach. The paper concludes by a brief discussion on model limitations and extensions.     

Novel (E)-3-(3-Oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides as Histone Deacetylase Inhibitors: Design,Synthesis and Bioevaluation

Herein, we report the design, synthesis and evaluation of novel (E)-3-(3-oxo-4-substituted-3,4-dihydro-2H-benzo[b][1,4]oxazin-6-yl)-N-hydroxypropenamides ( 4 a – i , 7 a – g ) targeting histone deacetylases. Three human cancer cell lines were used to test the cytotoxicity of the synthesized compounds (SW620, colon; PC-3, prostate; NCI−H23, lung cancer); inhibitory activity towards HDAC; anticancer activity; as well as their impact on the cell cycle and apoptosis. As a result, compounds 4 a – i bearing the alkyl substituents seemed to be less potent than the benzyl-containing compounds 7 a – g in all biological assays. Compounds 7 e – f were found to be the most active HDAC inhibitors with IC₅₀ of 1.498±0.020 μM and 1.794±0.159 μM, respectively. In terms of cytotoxicity and anticancer assay, 7 e and 7 f also showed good activity with IC₅₀ values in the micromolar range. In addition, the cell cycle and apoptosis of SW620 were affected by compound 7 f in almost a similar manner to that of reference compound SAHA. Docking assays were carried out for analysis the binding mode and selectivity of this compound toward 8 HDAC isoforms. Overall, our data confirmed that the inhibition of HDAC plays a pivotal role in their anticancer activity.     

CHAPS solubilization of a G-protein sensitive 5-HT1A receptor from bovine hippocampus

The binding of [3H] 8-OH-DPAT to membrane-bound 5-HT1A receptors from bovine hippocampus was saturable and corresponded to a single high-affinity state. Solubilization of the bovine hippocampal membranes with 10 mM CHAPS containing 200 mM NaCl, renders a preparation which binds [3H] 8-OH-DPAT with high-affinity (Kd = 1.9 nM) and is guanine nucleotide sensitive and ketanserin insensitive. 50% of [3H] 8-OH-DPAT binding activity is solubilized. The presence of GMP-P(NH)P promotes a low-affinity (Kd = 9.6 nM) state which is characteristic of receptors coupled to G-proteins. GMP-P(NH)P markedly accelerates the dissociation [3H] 8-OH-DPAT from solubilized membranes while having negligible effects on association. Thus, the agonist can activate the terniary complex rather than to promote its formation. 8-OH DPAT, WB 4101 and 5-carboxamidotryptamine dose responsively inhibit soluble [3H] 8-OH-DPAT binding with IC(50) values of 16.1, 15.6 and 1.3 nM, respectively. The CHAPS solubilized membrane preparation retains many of the [3H] 8-OH-DPAT binding characteristics of the membrane bound form.     

Object-oriented graphical modeling of FMSs

Presented in the article is a method for constructing a graphical model of an FMS by using a new modeling tool called JR-net (Job Resource relation-net). JR-net is an object-oriented graphical tool for modeling automated manufacturing systems (AMSs), such as FMSs, FASs, and AS/RSs. As with the object-oriented modeling paradigm of Rumbaugh et al. (1991), the JR-net modeling framework supports the three stages of models: static layout model (object model); job flow model (functional model); and supervisory control model (dynamic model). In this article, the existing JR-net structure (Park 1992, Han et al., 1995) is extended further to make it a graphical tool for FMS modeling. Using the extended JR-net, a step-by-step procedure for constructing a graphical model of FMSs is presented. Also addressed are issues of classifying FMSs in terms of their generic functions and of utilizing the JR-net model of FMSs.     

Enhancement of CO2 tolerance of Chlorella vulgaris by gradual increase of CO2 concentration

Summary Chlorella vulgaris UTEX259 was cultivated using two different methods of gas supply. In one method the CO₂ concentration in bubbled gas was held constant and in the other method it was increased gradually. Algal growth was almost linear after a short period of lag phase in both methods. With the constant CO₂ concentration, the CO₂ fixation rate in the linear growth phase decreased over 10%(v/v) CO₂, while the rate increased up to 6% CO₂. However, the rate was enhanced by using the latter incremental increase method, especially under a higher concentration of CO₂. The maximum rate of CO₂ fixation was 52 mg CO₂/l·h at 20% CO₂ during the gradual increase of CO₂ concentration.