Development of a Reliable Method to Express and Purify Vaccinia Virus Transcription Factors A7 and D6 in Bacteria

Poxvirus early genes code for viral products involved in host interaction, immune evasion, replication, and transcription of the viral genome. These viral early genes contain a unique and conserved promoter sequence that is recognized by early transcription factors (ETFs). The working hypothesis is pharmacologically targeting the structure of the poxvirus early promoters can inhibit transcription of poxvirus early genes. Our group has identified unique secondary structure which deviates from a scrambled control near the Vaccinia Virus (VACV) E9L promoter. Targeting these unique structures, which most likely may be G-quadruplexes, with structure specific ligands may distort local DNA structures at the promoter. In turn, this may affect the ability of viral transcription factors to recognize the target sequence. As the E9L gene codes for the viral DNA polymerase, inhibition of E9L transcription would prevent replication of the VACV DNA during infection. To assess potential antiviral activity of G-quadruplex ligands, the central goal of this thesis is to develop an in-vitro­ system to identify compounds that prevent binding of ETFs to target promoters. Using VACV ETFs (VETFs) as a model system, VETFs, A7L and D6R, were cloned into a bacterial expression vector with a His epitope tag for purification. A7 and D6 have successfully been expressed and detected in bacterial cell lysates via immunoblotting with anti-His antibodies. Currently, protocols are being optimized for large-scale purification of viral proteins to use in an in vitro binding assay

AN ANALYSIS OF 500 M INLINE SKATE STARTING MOTIONS

The purpose of this study was to examine if there are kinematical variables differences between national representative players (NRP) and non national representative players (NNRP) during 500 m inline skate starting motion. Four NRP and six NNRP were recruited for the study. Each subject executed starting motion five times on a 2x12 m start way in a gymnasium. Kinematical variables were analyzed by the three-dimensional motion analysis system (60Hz). It was hypothesized that there are time and center of mass acceleration differences in starting phase between groups since starting phase has been considered important in sprinting. The results showed that the NRP had significantly shorter starting phase time than that of NNRP

HOW CAN WE TEACH STUDENT TO ESTIMATE VERTICAL JUMP HEIGHTS USING GROUND REACTION FORCE DATA

The purpose of this study was to estimate vertical jump heights using ground reaction force (GRF) data and to suggest one practical example of biomechanical theory application to a real human motion. Vertical jump heights of impulse and flight time method were statistically smaller than three-dimensional video method. The causes of height differences seemed mainly from the fact that impulse was used to move jumper into the horizontal direction as well as into the vertical direction. Other important factors for accurate height calculation are jumper's mass and threshold value of GRF data collection. Vertical jump height calculation with GRF data showed an example of practical application of biomechanical theory to human motion and demonstrated a way of GRF equipment use for effective biomechanical theory education

The effect of palonosetron on rocuronium-induced withdrawal movement

AbstractBackgroundRocuronium causes pain and withdrawal movement during induction of anesthesia. In this study, palonosetron was investigated to have analgesic effect on the reduction of rocuronium-induced withdrawal movement.Methods120 patients were randomly assigned to one of three groups to receive either saline, lidocaine 20mg, or palonosetron 0.075mg with a tourniquet applied two minutes before thiopental sodium (5mg·kg−1) was given intravenously. After loss of consciousness, rocuronium (0.6mg·kg−1) was injected and the withdrawal movement was estimated by 4-point scale in a double-blind manner.ResultsThe overall incidence of rocuronium withdrawal movement was 50% with lidocaine (p=0.038), 38% with palonosetron (p=0.006) compared with 75% for saline. The incidence of no pain to mild pain was significantly lower in the lidocaine and palonosetron groups (85% and 92% respectively) than in the saline group (58%). However, there was no significant difference in withdrawal movement between the lidocaine and palonosetron groups. There was no severe movement with palonosetron.ConclusionPretreatment of palonosetron with venous occlusion may attenuate rocuronium-induced withdrawal movement as effective as the use of lidocaine. It suggested that peripheral action of palonosetron was effective to reduce rocuronium-induced withdrawal movement

Factors affecting work-related non-fatal injuries among aged workers in South Korea

IntroductionThe objective of this paper is to investigate whether an aging workforce is associated with an increase in work-related non-fatal injuries and to explore the underlying reasons for this potential increase.MethodsAged workers were defined as those who were at least 55-years-old. Work-related non-fatal injuries were assessed in aged and young workers who were registered with the workers’ compensation system from 2017 to 2021 of South Korea.ResultsThe mean estimated rate of work-related non-fatal injuries of aged workers (0.88/100) was about 2.5-times higher than that of younger workers (0.35/100). Most work-related non-fatal injuries in the older adults were in individuals working in the “construction sector” (36.0%), those with “elementary occupations (unskilled workers)” (45.0%), and those with employment status of “daily worker” (44.0%). “Trip & slip” (28.7%) and “falling” (19.6%) were more frequent types of work-related non-fatal injuries in aged workers relative to young workers. The category of “buildings, structures, and surfaces” was a more frequent cause of work-related non-fatal injuries in aged workers than young workers.DiscussionThe incidence of non-fatal work-related injuries is higher among aged workers compared to their younger counterparts. The increased occurrence of aged workers participating in precarious employment and jobs, along with the greater physical vulnerability, is likely the cause of their higher rate of work-related non-fatal injuries

Brd4 binds to active enhancers to control cell identity gene induction in adipogenesis and myogenesis

The epigenomic reader Brd4 is an important drug target for cancers. However, its role in cell differentiation and animal development remains largely unclear. Using two conditional knockout mouse strains and derived cells, we demonstrate that Brd4 controls cell identity gene induction and is essential for adipogenesis and myogenesis. Brd4 co-localizes with lineage-determining transcription factors (LDTFs) on active enhancers during differentiation. LDTFs coordinate with H3K4 mono-methyltransferases MLL3/MLL4 (KMT2C/KMT2D) and H3K27 acetyltransferases CBP/p300 to recruit Brd4 to enhancers activated during differentiation. Brd4 deletion prevents the enrichment of Mediator and RNA polymerase II transcription machinery, but not that of LDTFs, MLL3/MLL4-mediated H3K4me1, and CBP/p300-mediated H3K27ac, on enhancers. Consequently, Brd4 deletion prevents enhancer RNA production, cell identity gene induction and cell differentiation. Interestingly, Brd4 is dispensable for maintaining cell identity genes in differentiated cells. These findings identify Brd4 as an enhancer epigenomic reader that links active enhancers with cell identity gene induction in differentiation

Identification of a novel locus C2 controlling canary yellow flesh color in watermelons

The flesh color of watermelon is an important trait that is determined by carotenoid composition and affects consumers’ fruit desirability. Although a complete dominant control by C locus (Cllcyb) for canary yellow flesh (CY) over red flesh has been reported, red and CY colors frequently appear as a mixed pattern in the same flesh (incomplete canary yellow, ICY) in F1 and inbred lines carrying dominant C alleles. Therefore, we examined the genetic control of the mixed color pattern in ICY using whole-genome resequencing of three ICY (ICY group) and three CY inbred lines (CY group), as well as genetic linkage mapping of an F2 population. The segregation pattern in 135 F2 plants indicated that CY is controlled by a single locus (named C2) dominant over ICY. The whole-genome resequencing of ICY and CY inbred lines revealed an ICY/CY-specific region of approximately 27.60–27.88 Mb on Chr. 2 that was polymorphic between the ICY and CY groups. Our genetic map, using nine cleaved amplified polymorphic sequence markers developed based on the single-nucleotide polymorphisms from the ICY/CY-specific region, confirmed that C2 is located on Chr. 2 and cosegregated with the marker (M7) derived from a non-synonymous single-nucleotide polymorphism of the pentatricopeptide repeat (PPR) gene (ClPPR, Cla97C02G039880). Additionally, 27 watermelon inbred lines of ICY, CY, and red flesh were evaluated using previously reported Cllcyb (C locus)-based markers and our C2 locus-linked ClPPR-based marker (M7). As a result, dominant alleles at the C2 locus were required to produce CY, in addition to dominant alleles at the C locus, while a recessive homozygous genotype at the C locus gave the red flesh irrespective of the genotype at the C2 locus. Using a ClPPR-based cleaved amplified polymorphic sequence developed in this study and Cllcyb-based markers, watermelon cultivars with CY, ICY, and red flesh could be successfully discerned, implying that the combined use of these markers will be efficient for marker-assisted selection of flesh color in watermelon breeding

Genetic Diversity in Watermelon Cultivars and Related Species Based on AFLPs and EST-SSRs

The genetic relationships among 27 watermelon cultigens (Citrullus lanatus var. lanatus) from different countries of origin and with different horticultural characteristics and 5 related wild-type species and subspecies (Citrullus colocynthis, Citrullus lanatus var. citroides, and Citrullus rehmii) were assessed using amplified fragment length polymorphism (AFLP) and expressed sequence tag-simple sequence repeat (EST-SSR) markers. AFLPs were evaluated using 16 EcoRI-MseI primer combinations, and 862 alleles (an average of 53.8 alleles per primer combination) were scored. Polymorphisms were found in 806 (93.4%) alleles, whereas 56 monomorphic alleles were identified. Using 16 EST-SSR primer sets, 103 alleles were scored, and all 103 alleles were polymorphic among the 32 genotypes with an average of 6.4 polymorphic alleles per primer pair. However, the high polymorphic ratio among the AFLPs and EST-SSRs was largely due to the wild-type species, while little diversity was observed among the adapted cultivars. Genetic similarity coefficients were calculated based on the 965 polymorphic AFLP and EST-SSR alleles, and a phenetic tree was constructed. The dendrogram contained 2 major clusters. Cluster I included all adapted watermelon cultivars, and the similarity among these cultigens was very high (0.94-0.98), demonstrating cross relationships and a narrow genetic background. Cluster II was composed of 4 wild-type species. The genetic distance between the nodes that comprise these 2 clusters was approximately 0.63, indicating a high level of genetic dissimilarity between the adapted watermelon species and the other related species. The low level of marker polymorphism among the adapted cultivars implies that a severe bottleneck in genetic diversity existed in watermelon during the initial breeding practices