RESUMEN
We have developed and tested two linked but separable structured inquiry exercises using a set of Drosophila melanogaster GAL4 enhancer trap strains for an upper-level undergraduate laboratory methods course at Bucknell University. In the first, students learn to perform inverse PCR to identify the genomic location of the GAL4 insertion, using FlyBase to identify flanking sequences and the primary literature to synthesize current knowledge regarding the nearest gene. In the second, we cross each GAL4 strain to a UAS-CD8-GFP reporter strain, and students perform whole mount CNS dissection, immunohistochemistry, confocal imaging, and analysis of developmental expression patterns. We have found these exercises to be very effective in teaching the uses and limitations of PCR and antibody-based techniques as well as critical reading of the primary literature and scientific writing. Students appreciate the opportunity to apply what they learn by generating novel data of use to the wider research community.
Asunto(s)
Curriculum , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Elementos de Facilitación Genéticos , Laboratorios , Aprendizaje , Factores de Transcripción/genética , Universidades , Animales , Secuencia de Bases , Encéfalo/metabolismo , Regulación de la Expresión Génica , Genes de Insecto , Datos de Secuencia Molecular , Cuerpos Pedunculados/metabolismo , Reacción en Cadena de la PolimerasaRESUMEN
Cell and gene therapies (CGTs) have revolutionized patient outcomes and provided care options for previously untreatable conditions. The clinical and commercial progress of CGT therapies is hindered by chemistry, manufacturing, and control (CMC) challenges. This article summarizes recommendations from the 2023 Annual Meeting CMC sessions wherein speakers advocated for science-driven comparability strategies, proactive risk assessments, clearer regulatory guidance, and a shift from retrospective to prospective studies. Planning for manufacturing changes, statistical approaches, and consideration of multiple product versions also emerged as crucial elements to help sponsors navigate CMC hurdles for successful CGT clinical and commercial development.
RESUMEN
Human cytomegalovirus (HCMV), a member of the Betaherpesvirinae sub-family of Herpesviridae family, is a widespread pathogen that infects a majority of the world's population by early adulthood. In individuals whose immune systems are immature or weakened, HCMV is a significant pathogen causing morbidity and mortality. There is no effective vaccine and only limited antiviral treatments against HCMV infection to date. A possible target for novel antiviral treatments is the HCMV proteins that localize to the tegument of the virion, since they play important roles in all stages of the viral life cycle, including, viral entry, gene expression, immune evasion, assembly, and egress. The most likely tegument protein candidates are pp65 (immune evasion), pp71 (gene expression), and pp150 and pp28 (assembly and egress). Although the subcellular localization of these proteins has been identified during HCMV infections in vitro, their localization patterns have not been determined when each protein is expressed individually in living cells. Thus, the objective of this review is elucidate the HCMV tegument as well as present current research findings concerning the subcellular localization of the tegument proteins pp65, pp71, pp150, and pp28 as fusions to one of several fluorescent proteins.