Identification of the Actin-Binding Region and Binding to Host Plant Apple Actin of Immunodominant Transmembrane Protein of ' Phytoplasma mali'.

' Phytoplasma mali' ('. P. mali') has only one major membrane protein, the immunodominant membrane protein (Imp), which is regarded as being close to the ancestor of all phytoplasma immunodominant membrane proteins.

Beyond Destabilizing Activity of SAP11-like Effector of Phytoplasma mali Strain PM19.

It was shown that the SAP11 effector of different Phytoplasma can destabilize some TEOSINE BRANCHES/CYCLOIDEA/PROLIFERATING CELL FACTORs (TCPs), resulting in plant phenotypes such as witches' broom and crinkled leaves. Some SAP11 exclusively localize in the nucleus, while the others localize in the cytoplasm and the nucleus. The SAP11-like effector of Phytoplasma mali strain PM19 (SAP11) localizes in both compartments of plant cells.

The Effect of the Anticipated Nuclear Localization Sequence of ' Phytoplasma mali' SAP11-like Protein on Localization of the Protein and Destabilization of TCP Transcription Factor.

SAP11 is an effector protein that has been identified in various phytoplasma species. It localizes in the plant nucleus and can bind and destabilize TEOSINE BRANCHES/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) transcription factors. Although SAP11 of different phytoplasma species share similar activities, their protein sequences differ greatly.

Bottom-up assembly of a bilayer structure of icosahedral viral nanoparticles.

The development of 2D and 3D structures on the nanoscale containing viral nanoparticles (VNPs) as interesting nanobuilding blocks has come into focus for a bottom-up approach as an alternative to the top-down approach in nanobiotechnology. Our research has focused on the plant Tomato Bushy Stunt Virus (TBSV). In a previous study, we reported the impact of the pH value on the 2D assembly of viral monolayers.

' Phytoplasma mali' Genome Encodes a Protein that Functions as an E3 Ubiquitin Ligase and Could Inhibit Plant Basal Defense.

Phytoplasmas are the causative agent of numerous diseases of plant species all over the world, including important food crops. The mode by which phytoplasmas multiply and behave in their host is poorly understood and often based on genomic data. We used yeast two-hybrid screening to find new protein-protein interactions between the causal agent of apple proliferation ' Phytoplasma mali' and its host plant.

Electrostatic conditions define the 2D self-assembly of tomato bushy stunt viruses on solid surfaces.

Plant viruses which are self-assembled on a substrate are interesting building blocks in nanobiotechnology, in particular, for the creation of 2D ordered structures. In this article, the self-assembly of different genetically modified types of the tomato bushy stunt virus spin-coated on pristine silicon was investigated by scanning force and scanning electron microscopy. Amino acid side chains were integrated in the capsids of the viruses by extending the coat protein with different charged amino acid clusters (tetra-aspartate-hexa-histidine, hexa-aspartate, or tetra-arginine-tags).

A simple method to estimate the isoelectric point of modified Tomato bushy stunt virus (TBSV) particles.

We present a simple method to estimate the isoelectric point (pI) of Tomato Bushy Stunt particles. We demonstrate that the combination of agarose gels with different pH buffers can be used to electrophorese the virus particles and their migration patterns can be compared. This method allows us to estimate the pI of the virus particles (wild type, wt, and genetically modified particles) and to monitor the effect of the pI of modified peptide side chains of the viral capsid subunit on the pI of the whole virus particle.

Identification of Highly Specific scFvs against Total Adiponectin for Diagnostic Purposes.

Adiponectin is one of the most abundant adipokines secreted from adipose tissue. It acts as an endogenous insulin sensitizer and plasma concentrations are inversely correlated with obesity and metabolic syndrome. A decrease in plasma adiponectin levels normally indicates increased hormonal activity of the visceral lipid tissue, which is associated with decreased insulin sensitivity.

Immunodominant membrane proteins of phytoplasmas.

Phytoplasmas are plant-pathogenic, phloem-colonizing, cell wall-less microorganisms that are primarily dependent on insect transmission for their spread and survival. The life cycle of phytoplasmas involves replication in insects and host plants. Until recently, phytoplasmas have resisted all attempts at cultivation in cell-free media, making these pathogens poorly characterized on a physiological and biochemical basis.

Binding and processing of small dsRNA molecules by the class 1 RNase III protein encoded by sweet potato chlorotic stunt virus.

Sweet potato chlorotic stunt virus (SPCSV; genus Crinivirus, family Closteroviridae) causes heavy yield losses in sweet potato plants co-infected with other viruses. The dsRNA-specific class 1 RNase III-like endoribonuclease (RNase3) encoded by SPCSV suppresses post-transcriptional gene silencing and eliminates antiviral defence in sweet potato plants in an endoribonuclease activity-dependent manner. RNase3 can cleave long dsRNA molecules, synthetic small interfering RNAs (siRNAs), and plant- and virus-derived siRNAs extracted from sweet potato plants.

Improvement of PCR reaction conditions for site-directed mutagenesis of big plasmids.

QuickChange mutagenesis is the method of choice for site-directed mutagenesis (SDM) of target sequences in a plasmid. It can be applied successfully to small plasmids (up to 10 kb). However, this method cannot efficiently mutate bigger plasmids.

An immunodominant membrane protein (Imp) of 'Candidatus Phytoplasma mali' binds to plant actin.

The phytopathogenic, cell-wall-less phytoplasmas exhibit a dual life cycle: they multiply in the phloem of their host plant and in the body of their insect vector. Their membrane proteins are in direct contact with both hosts and are supposed to play a crucial role in the phytoplasma spread within the plant as well as by the insect vector. Three types of nonhomologous but highly abundant and immunodominant membrane proteins (IDP) have been identified within the phytoplasmas: Amp, IdpA, and Imp.

Tomato bushy stunt viruses (TBSV) in nanotechnology investigated by scanning force and scanning electron microscopy.

Spherical plant viruses like the tomato bushy stunt virus (TBSV) allow for multiple applications in nanotechnology due to their shape. In this article, different types of the virus were created by extending coat protein (CP) at carboxylic termini with 2 different charged amino acids by point mutation. The obtained CPs carried 6 aspartic acid (negative charge) and 4 histamine (positive charge) residues.

Analysis of the autoproteolytic activity of the recombinant helper component proteinase from zucchini yellow mosaic virus.

The multifunctional helper component proteinase (HC-Pro) of potyviruses contains an autoproteolytic function that, together with the protein 1 (P1) and NIa proteinase, processes the polyprotein into mature proteins. In this study, we analysed the autoproteolytic active domain of zucchini yellow mosaic virus (ZYMV) HC-Pro. Several Escherichia coli-expressed MBP:HC-Pro:GFP mutants containing deletions or point mutations at either the N- or C-terminus of the HC-Pro protein were examined.

Diverse spontaneous silencing of a transgene among two Nicotiana species.

In plants, transgenes frequently become spontaneously silenced for unknown reasons. Typically, transgene silencing involves the generation of small interfering RNAs (siRNAs) that directly or indirectly target cognate DNA and mRNA sequences for methylation and degradation, respectively. In this report, we compared spontaneous silencing of a transgene in Nicotiana benthamiana and Nicotiana tabacum.

The helper component-proteinase of the Zucchini yellow mosaic virus inhibits the Hua Enhancer 1 methyltransferase activity in vitro.

The helper component-proteinase (HC-Pro) is a multifunctional protein found among potyviruses. With respect to its silencing suppressor function, small RNA binding appears to be the major activity of HC-Pro. HC-Pro could also exhibit other suppressor activities.

Expression, purification and functional characterization of recombinant Zucchini yellow mosaic virus HC-Pro.

HC-Pro is a helper component-proteinase which acts as a multifunctional protein in the potyviral life cycle. Apart from its proteolytic activity, HC-Pro has the capacity to bind duplex small RNAs (sRNAs). To investigate HC-Pro-mediated sRNA binding in vitro, high amounts of purified protein are required.

First description of Cryptosporidium bovis in Japan and diagnosis and genotyping of Cryptosporidium spp. in diarrheic pre-weaned calves in Hokkaido.

Eighty fecal samples from pre-weaned calves with diarrhea were collected in the Tokachi area in Northern Japan to investigate the prevalence of Cryptosporidium species in such animals. Oocysts from fecal samples collected from each animal were concentrated using sucrose gradient centrifugation. Genomic DNA was extracted from each sample and processed by nested PCR to amplify the partial SSU rRNA gene of Cryptosporidium.

Analysis of an autoproteolytic activity of rice yellow mottle virus silencing suppressor P1.

Ectopically expressed rice yellow mottle virus P1 fusion proteins were found to be cleaved in planta and in Escherichia coli. Cleavage takes place in the absence of bacterial protease activity, indicating that the P1 fusion is autocatalytically processed independently of host factors. N-terminal sequencing of the C-terminal cleavage product of transiently expressed P1/GFP (green fluorescence protein) in Nicotiana benthamiana showed that the cleavage site is located between the first two amino acids (aa) downstream of the P1 sequence.

Analysis of the effectiveness of reused primary and secondary antibodies in Western blotting analysis.

Western blot analysis is a common technique for detecting a target protein on a nitrocellulose membrane. The target protein is detected using specific antibodies. The positive signal can be visualized either by chemiluminescence or by a colorimetric reaction.

Homemade site directed mutagenesis of whole plasmids.

Site directed mutagenesis of whole plasmids is a simple way to create slightly different variations of an original plasmid. With this method the cloned target gene can be altered by substitution, deletion or insertion of a few bases directly into a plasmid. It works by simply amplifying the whole plasmid, in a non PCR-based thermocycling reaction.

Association of low CD4 cell count and intrauterine growth retardation in Thailand.

Objective: Each year, intrauterine growth retardation (IUGR) affects 20-30 million neonates worldwide, mostly in resource-limited settings. Increased perinatal and infant mortality has been associated with IUGR. Some studies have suggested that HIV infection could increase the risk of IUGR.

Prevalence and molecular characterization of human and bovine Cryptosporidium isolates in Thailand.

This study was performed to determine the prevalence and genotypes of Cryptosporidium species among HIV patients and cattle in Thailand. Stool specimens were collected from 46 HIV patients from Prabat Nampu Temple, Lop Buri Province in central Thailand. Two hundred fecal samples from dairy cattle were collected from seven farms in Chon Buri Province, the eastern part of Thailand.

Efficient procedure for grapevine embryogenic suspension establishment and plant regeneration: role of conditioned medium for cell proliferation.

An efficient system for the establishment and multiplication of highly prolific embryogenic cell cultures of grapevine (Vitis sp.) was developed. Using anther-derived pro-embryogenic masses as starting material, cell suspensions of different grapevine cultivars (Tempranillo, Cabernet-Sauvignon) and rootstocks (Kober 125 AA, Kober 5 BB, 110 Richter) were initiated in liquid medium containing NOA (1.