Accumulated data and results from the recent study of dsRNA isolated from grapevines used in experiments of insect and graft transmission of 'Shiraz' disease

RT-PCR amplicons of dsRNA isolated from various grapevines, which were used in the experiments of transmission of 'Shiraz' disease (SD) from 'Cinsaut Blanc' clone P163/12 to SD-susceptible 'Merlot' and 'Shiraz' using mealybug Planococcus ficus and grafting were investigated. The amplicons were generated in RT-PCR based on virus-specific or random hexamers oligonucleotide primers. Standard molecular techniques and high-throughput sequencing (HTS), respectively, were applied. The results supported the hypothesis that GVA M5v variant present in 'Cinsaut Blanc' P163/12, which is a member of group II of GVA variants associated with SD, is crucial for developing this disease. HTS data did not reveal any other grapevine viruses besides GLRaV-3 and GVA in SD-affected grapevines, except for GVE which, however, was not present in all diseased plants

Alpha, Beta, Launch: A Newbie\u27s Guide to Educational Video Game Development

This paper details the process we went through to develop an educational video game, which includes: research on implementing video games into the classroom, vendor selection, video game design, and curriculum development. Throughout the video game development process, we faced challenges such as budget, time constraint, and varying areas of expertise. This paper serves as a guideline for similar organizations interested in educational video game development. Play game on desktop or tablet: www.avma.org/videogame Play within browser: https://www.avma.org/KB/K12/videogame/index.htm

Single-strand conformation polymorphism (SSCP), cloning and sequencing reveals two major groups of divergent molecular variants of grapevine leafroll-associated virus 3 (GLRaV-3)

The SSCP and RE/SSCP analysis of ORF5, ORF6 and ORF7 of 25 local and overseas isolates of GLRaV-3 showed only two kinds of distinct SSCP profiles for each of these genomic regions. It suggested low molecular variability of the virus. Fragments of the 5’UTR + ORF1a, 3’ terminal part of ORF1a, ORF4, ORF5, ORF6 and ORF7 of three isolates, representing distinct SSCP profiles, were cloned and sequenced. Results revealed that although the 3’terminal half of the genome (ORF4-7) and the sequence located in the 3’ terminal part of ORF1a were relatively similar among isolates (91.3-96.2 % nt identity), their 5’terminal parts (88 nt of 5’UTR and adjacent first 329 nt of ORF1a) were clearly divergent (81.6-81.8 % nt identity). Analysis of this divergent part of GLRaV-3 for an additional 11 isolates showed that they cluster in two distinct molecular groups, sharing 94.7-99.7 % and 80.8-85.1 % nt identity within and between groups respectively. The correlation between the molecular groups and SSCP profiles of the 209 nt fragment of ORF5 of GLRaV-3 strongly suggests that SSCP analysis of this easily RT-PCR amplified region can be used for rapid identification of divergent molecular variants of the virus in field-collected grapevine samples.

The application of single-strand conformation polymorphism (SSCP) technique for the analysis of molecular heterogeneity of grapevine virus A

The results of the analysis of grapevine virus A (GVA) isolates by single-strand conformation polymorphism (SSCP) confirm that this technique is very helpful in rapid and relatively low cost preliminary analysis of molecular heterogeneity of viruses. The results clearly show that the reliability of SSCP analysis of GVA depends on oligonucleotide primers for successful RT-PCR amplification of the highest possible number of molecular variants of the virus. Among 7 pairs of GVA-specific primers designed in different laboratories only two, those from Canada (117038 and C7273) and Switzerland (MP and CPdt), allowed positive RT-PCR amplification of all our isolates of the virus mechanically transmitted from various grapevines to Nicotiana benthamiana. With SSCP analysis of 238 bp DNA fragments complementary to part of ORF5 of GVA, produced by RT-PCR using the first pair of primers, we were able to detect 1-35 nt differences between GVA isolates. The DNA fragments, about 986 bp, complementary to part of ORF3 and ORF4, ORF5 and 3'UTR of GVA, produced by RT-PCR using the second pair of primers, were useful for SSCP analysis only after their digestion with the restriction enzyme DdeI. The results strongly suggest that SSCP analysis of 238 nt fragment of ORF5 of GVA along with DdeI/SSCP analysis of about 986 nt 3'terminal fragment of the virus allow rapid and reliable determination of the number of dominant nt sequence variants of GVA present in a single N. benthamiana or grapevine plant.

Genome Sequences and Structures of Two Biologically Distinct Strains of Grapevine leafroll - associated virus 2 and Sequence Analysis

Grapevine leafroll-associated virus 2 (GLRaV-2), a member of the genus Closterovirus within Closteroviridae, is implicated in several important diseases of grapevines including "leafroll”, "graft-incompatibility”, and "quick decline” worldwide. Several GLRaV-2 isolates have been detected from different grapevine genotypes. However, the genomes of these isolates were not sequenced or only partially sequenced. Consequently, the relationship of these viral isolates at the molecular level has not been determined. Here, we group the various GLRaV-2 isolates into four strains based on their coat protein gene sequences. We show that isolates "PN” (originated from Vitis vinifera cv. "Pinot noir”), "Sem” (from V. vinifera cv. "Semillon”) and "94/970” (from V. vinifera cv. "Muscat of Alexandria”) belong to the same strain, "93/955” (from hybrid "LN-33”) and "H4” (from V. rupestris "St. George”) each represents a distinct strain, while Grapevine rootstock stem lesion-associated viru

Creating chimeras: Embryonic stem cells incorporated

Gene modification within the murine genome has become a powerful and invaluable tool to investigate development. One example of the utility of such technology is providing a method by which researchers can follow cells throughout development via the introduced genetic modifications.Fil: Pascottini, Osvaldo Bogado. University of Chicago; Estados UnidosFil: Goszczynski, Daniel Estanislao. University of Chicago; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Nguyen, Alexandra L.. University of Chicago; Estados Unido

Caracterización de polimorfismos en los genes PPARG, CEBPA, LIPE, RXRA y FABP4 asociados a metabolismo lipídico en razas de ganado bovino

La calidad de la carne está determinada por cualidades como el marmoleo, el sabor, la terneza y la composición, entre otras. Estas cualidades están reguladas a distintos niveles, y uno de ellos es la genética. Hoy en día se conoce buena parte de las vías metabólicas que regulan estas características, y se han propuesto "genes candidatos" que codifican factores importantes dentro de estas vías. Los genes PPARG, CEBPA, FABP4, LIPE y RXRA son parte de las vías de diferenciación adipocítica y del metabolismo lipídico. El objetivo de este proyecto fue caracterizar la variabilidad genética en estos genes en razas bovinas con diferente calidad carnicera. Los datos se obtuvieron por medio de técnicas moleculares (reacción en cadena de la polimerasa, re-secuenciación) aplicadas a muestras de ADN extraídas de animales pertenecientes a diferentes razas criadas alrededor del mundo. Luego se realizaron una serie de análisis a través de programas bioinformáticos y herramientas web. Algunos de los polimorfismos detectados en los genes y otros disponibles en las bases de datos de internet fueron seleccionados para realizar estudios de validación a nivel poblacional y análisis estadísticos de asociación a caracteres de calidad carnicera en una población de ganado local. Los resultados fueron diversos: PPARG y CEBPA presentaron una variabilidad moderada, y FABP4 y LIPE presentaron una variabilidad alta. Algunos de los polimorfismos analizados sugieren una asociación a la composición lipídica de la carne y otros caracteres de engrasamiento, como espesor de grasa dorsal. Algunas de las posibles explicaciones biológicas para estas asociaciones fueron analizadas con diferentes herramientas bioinformáticas y se observaron algunos fenómenos interesantes. El conocimiento de la variabilidad existente en estos genes es de importancia para complementar los métodos de selección genética tradicionales y mejorar la calidad del ganado.Facultad de Ciencias Veterinaria

Production and use of an antiserum to grapevine virus B capsid protein purified from SDS-polyacrylamide gels

Antiserum to electrophoretically-separated capsid protein of grapevine virus B (GVB) was produced. After easy and effective elimination of antibodies cross-reactive with grapevine virus A (GVA), the antiserum was successfully used in ELISA for the detection of GVB in grapevines

Capturing chaotic chromosomes: Pairing in action

Accurate chromosome segregation is critical for the formation of haploid gametes, and therefore healthy offspring. Errors in segregation result in aneuploidy, which increases exponentially with maternal age (Hassold and Hunt, 2001). Maternally aged mouse oocytes were recently found to exhibit premature sister chromatid separation due to reduced levels of Securin, an essential regulator of sister chromatid cohesion (Nabti et al, 2017). This landmark finding was facilitated by using chromosome spreads that allow for the visualization of meiotic processes, such as recombination, synapsis, crossing over, and cohesion. This powerful technique is easy to perform and analyze, and allows for the identification of markers for different processes, including DNA damage and repair.Fil: Bertucci, Micka C.. University of Chicago; Estados UnidosFil: Das, Arunika. University of Chicago; Estados UnidosFil: Fitzgerald, Harriet C.. University of Chicago; Estados UnidosFil: Goszczynski, Daniel Estanislao. University of Chicago; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Detection of two strains of grapevine leafroll-associated virus 2

Two strains of grapevine leafroll-associated virus 2 (GLRaV-2) were obtained by mechanical transmission from grapevines to Nicotiana benthantiana. The strains, designated 94/970 and 93/955, consistently differed with regard to the development of symptoms. The first induced chlorotic and occasional white-necrotic local lesions while the second induced chlorotic followed by metallic-opalescent, solid necrotic local lesions. The strains were indistinguishable with regard to the molecular weight of their capsid proteins or serologically. A difference in the pattern of minor dsRNA bands was consistently observed