Molecular and statistical approaches to the detection and correction of errors in genotype databases.

Errors in genotyping data have been shown to have a significant effect on the estimation of recombination fractions in high-resolution genetic maps. Previous estimates of errors in existing databases have been limited to the analysis of relatively few markers and have suggested rates in the range 0.5%-1.5%. The present study capitalizes on the fact that within the Centre d'Etude du Polymorphisme Humain (CEPH) collection of reference families, 21 individuals are members of more than one family, with separate DNA samples provided by CEPH for each appearance of these individuals. By comparing the genotypes of these individuals in each of the families in which they occur, an estimated error rate of 1.4% was calculated for all loci in the version 4.0 CEPH database. Removing those individuals who were clearly identified by CEPH as appearing in more than one family resulted in a 3.0% error rate for the remaining samples, suggesting that some error checking of the identified repeated individuals may occur prior to data submission. An error rate of 3.0% for version 4.0 data was also obtained for four chromosome 5 markers that were retyped through the entire CEPH collection. The effects of these errors on a multipoint map were significant, with a total sex-averaged length of 36.09 cM with the errors, and 19.47 cM with the errors corrected. Several statistical approaches to detect and allow for errors during linkage analysis are presented. One method, which identified families containing possible errors on the basis of the impact on the maximum lod score, showed particular promise, especially when combined with the limited retyping of the identified families. The impact of the demonstrated error rate in an established genotype database on high-resolution mapping is significant, raising the question of the overall value of incorporating such existing data into new genetic maps.     

Study of mechanisms of electric field-induced DNA transfection. V. Effects of DNA topology on surface binding, cell uptake, expression, and integration into host chromosomes of DNA in the mammalian cell.

Neumann and coworkers (Neumann, E., M. Schaefer-Ridder, Y. Wang, and P. H. Hofschneider. 1982. EMBO J. 1:841-845) have shown that the efficiency of pulsed electric field (PEF)-induced DNA transfection of mouse L-cells by the thymidine kinase gene is several times higher for the linear DNA than for the closed circular DNA. Transfection of Escherichia coli bacteria by several plasmids indicates that the transfection efficiency was much higher for the closed circular/supercoiled (sc-) and circular/relaxed (cr-) DNA than for the linearized (In-) DNA (Xie, T. D., L. Sun, H. G. Zhao, J. A. Fuchs, and T. Y. Tsong. 1992. Biophys. J. 63:1026-1031). To resolve these conflicting observations, we have systematically examined electrotransfection of NIH3T3 mouse fibroblast by the plasmids, pRSVcat, pRSVneo, and pRSVgpt. Mg(2+)-facilitated surface binding of DNA before, and DNA uptake by 3T3 cells after treatment with PEF, were monitored by 3H-labeled plasmids. Transfection efficiency was evaluated by both the transient expression of chloramphenicol acetyltransferase (cat) activity 2-3 days after, and the permanent expression of neomycin phosphotransferase (neo) and xanthine-guanine phosphoribosyltransferase (gpt) genes in the transformants 2 weeks after the PEF treatment. Our results indicate that cell surface binding and PEF-induced cell uptake of DNA did not depend on the topology of DNA. However, both the transient and the permanent expression of the plasmids were three to five times more efficient for the cr-DNA and the sc-DNA than for the in-DNA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Light-regulated and cell-specific expression of tomato rbcS-gusA and rice rbcS-gusA fusion genes in transgenic rice.

A previously isolated rice (Oryza sativa) rbcS gene was further characterized. This analysis revealed specific sequences in the 5' regulatory region of the rice rbcS gene that are conserved in rbcS genes of other monocotyledonous species. In transgenic rice plants, we examined the expression of the beta-glucuronidase (gusA) reporter gene directed by the 2.8-kb promoter region of the rice rbcS gene. To examine differences in the regulation of monocotyledonous and dicotyledonous rbcS promoters, the activity of a tomato rbcS promoter was also investigated in transgenic rice plants. Our results indicated that both rice and tomato rbcS promoters confer mesophyll-specific expression of the gusA reporter gene in transgenic rice plants and that this expression is induced by light. However, the expression level of the rice rbcS-gusA gene was higher than that of the tomato rbcS-gusA gene, suggesting the presence of quantitative differences in the activity of these particular monocotyledonous and dicotyledonous rbcS promoters in transgenic rice. Histochemical analysis of rbcS-gusA gene expression showed that the observed light induction was only found in mesophyll cells. Furthermore, it was demonstrated that the light regulation of rice rbcS-gusA gene expression was primarily at the level of mRNA accumulation. We show that the rice rbcS gene promoter should be useful for expression of agronomically important genes for genetic engineering of monocotyledonous species.     

血清岩藻糖测定方法学研究

本文对血清岩藻糖测定方法进行了系统研究。血清用量由200μL减至50μL,显色反应4h内稳定。吸收峰在396nm。岩藻糖浓度40μg/mL内线性良好,批间CV=2.1％。以Sephadex-G200层析,血清岩藻糖主要存在于分子量为500kD的组分中。以本法测得30例健康人血清岩藻糖浓度为588.1±172.0μmol/L。     

红毛五加多糖对腹腔巨噬细胞作用的定量细胞化学研究

中药红毛五加(Acanthopanax giraldii Harms)属五加科植物,本文通过细胞化学定性、定位、定量研究探讨红毛五加多糖(AGPS)对腹腔巨噬细胞的作用及机理。实验证明AGPS能使巨噬细胞数量明显增多,细胞体积增大,伪足增多,吞噬能力增强,细胞内醣类、酸性磷酸酶、三磷酸腺昔酶、酸性酯酶和琥珀酸脱氢酶活性显著增强。用显微分光光度计对上述单个细胞的化学成分进行定量测定。实验组和对照组结果有显著差异。提示红毛五加的扶正固本作用十分明显,本研究为AGPS的应用和作用机理提供了一定的实验依据。     

G418对IL—2基因包装细胞的筛选

本文用高浓度的G418(800μg/ml)和低浓度的G418(200μg/ml)对包装细胞PLXSN/IL-2/PA317细胞进行40天的选择筛选培养,使其细胞呈稳定状态生长时,收集上清液转染NIH/3T3细胞,进行病毒滴度测定。试图在高选择标记的情况下筛选出高表达目的基因的包装细胞。实验结果表明:高、低浓度的G418对PLXSN/IL-2/PA317包装细胞的选择作用相同,即包装细胞的病毒滴度同选择标记物浓度无关。提示可用低浓度的G418来维持包装细胞的生命。     

Three research priorities for just and sustainable urban systems: Now is the time to refocus

Now is the time to refocus efforts in urban research and design. A changing climate and extreme weather events are presenting unique challenges to urban systems around the world. These challenges illuminate the social barriers that accompany disruptive events such as resource inequities and injustices. In this perspective, we provide three research priorities for just and sustainable urban systems that help to address these matters. The three research priorities are: (1) social equity and justice, (2) circularity, and (3) digital twins. Conceptual context and future research directions are provided for each. For social equity and justice, the future directions are mandatory equity analysis and inclusionary practices, understanding and reconciling historical injustices, and intentional integration with diverse community stakeholders. For circularity applications, they are better metrics for integration, more robust evaluation frameworks, and dynamic modeling at multiple spatial and temporal scales. Future directions for digital twins include developing principles to reduce complexity, integrating model and system components, and reducing barriers to data access. These research priorities are core to meeting several of the United Nations Sustainable Development Goals (i.e., 1—No Poverty, 8—Decent Work and Economic Growth, 10—Reduced Inequalities, and 11—Sustainable Cities and Communities). Useful social and technical matters are discussed throughout, where we highlight the importance of prioritizing localized research efforts, provide guidance for community-engaged research and co-development practices, and explain how these priorities interact to align with the evolving field of industrial ecology.     

Circular RNA SLTM as a miR-421-competing endogenous RNA to mediate HMGB2 expression stimulates apoptosis and inflammation in arthritic chondrocytes

Osteoarthritis (OA) is the most common age-related joint disease characterized by chronic inflammation, progressive articular cartilage destruction, and subchondral sclerosis. Accumulating evidence suggests that circular RNAs (circRNAs) play key roles in OA, but the function of circSLTM in OA remains greatly unknown. Therefore, this study focused on interleukin-1β (IL-1β)-treated primary human chondrocytes as well as a rat model to investigate the expression pattern and functional role of circSLTM in OA in vitro and in vivo. CircSLTM and high mobility group protein B2 (HMGB2) were upregulated in IL-1β-induced chondrocytes, whereas miR-421 was downregulated. Knockdown of circSLTM or overexpression of miR-421 ameliorated IL-1β-induced chondrocyte apoptosis and inflammation. The regulatory relationship between circSLTM and miR-421, as well as that between miR-421 and HMGB2, was predicted by bioinformatics and then verified by the RNA immunoprecipitation experiment and dual-luciferase reporter gene assay. Furthermore, silencing of circSLTM increased cartilage destruction and decreased cartilage tissue apoptosis rate and inflammation in a rat model of OA. Taken together, our findings demonstrate the fundamental role of circSLTM in OA progression and provide a potential molecular target for OA therapy.