Efficient cDNA cloning by direct phenotypic correction of a mutant human cell line (HPRT-) using an Epstein-Barr virus derived cDNA expression vector.

Human cells are, in general, poor recipients of foreign DNA, which has severely hampered the cloning of genes by direct phenotypic correction of deficient human cell lines after DNA mediated gene transfer. In this communication a methodology is presented which largely circumvents this problems. The method relies on the use of a recently developed episomal Epstein-Barr-virus-derived cDNA expression vector (Belt et al. (1989) Gene 84, 407-417). The cloning of hypoxanthine phosphoribosyltransferase (HPRT) cDNA, corresponding to a low abundant mRNA in wild type cells is used as a model system. Size fractionated poly (A)+ RNA from wild type cells, which resulted in an approximately 10 fold enrichment in HPRT mRNA, was used to construct a cDNA library of 25,000 independent clones in the pECV25 vector. An HPRT deficient human cell line was transfected and subsequently selected with hygromycin B for DNA uptake. In a small scale experiment only 7000 hygromycin BR transfectants were sufficient to isolate 2 independent HATR clones which were shown to replicate episomes harbouring HPRT cDNA. The first insert had a 5' untranslated region (UTR) and a 3' UTR perfectly in agreement with published data. The second cDNA clone harboured an unusually long 5' UTR and a shorter 3' UTR due to alternative polyadenylation of the HPRT transcript which has not been previously recognized

Partially distributed coordination with Reo and constraint automata

Algorithms and the Foundations of Software technolog

Service orchestration with priority constraints

Business process management is an operational management approach that focuses on improving business processes. Business processes, i.e., collections of important activities in an organization, are represented in the form of a workflow, an orchestrated and repeatable pattern of activities amenable to automated analysis and control. Priority is an important concept in modeling workflows. We need priority to model cancelable and compensable tasks within transactional business processes. We use the Reo coordination language to model and formally analyze workflows. In this paper, we propose a constraint-based approach to formalize priority in Reo. We introduce special channels to propagate and block priority flows, define their semantics as constraints, and model priority propagation as a constraint satisfaction problem

Algebraic service composition for user-centric IoT applications

The Internet of Things (IoT) requires a shift in our way of building applications, as it is aimed at providing many services to society in general. Non-developer people require increasingly complex IoT applications and support for their ever changing run-time requirements. Although service composition allows the combination of functionality into more complex behaviours, current approaches provide support for dealing with one IoT scenario at a time, as they allow the definition of only one workflow. In this paper, we present DX-MAN, an algebraic model for static service composition that allows the definition of composite services that encompass multiple workflows for run-time scenarios. We evaluate our proposal on an example in the domain of smart homes

Use of specific Green's functions for solving direct problems involving a heterogeneous rigid frame porous medium slab solicited by acoustic waves

A domain integral method employing a specific Green's function (i.e., incorporating some features of the global problem of wave propagation in an inhomogeneous medium) is developed for solving direct and inverse scattering problems relative to slab-like macroscopically inhomogeneous porous obstacles. It is shown how to numerically solve such problems, involving both spatially-varying density and compressibility, by means of an iterative scheme initialized with a Born approximation. A numerical solution is obtained for a canonical problem involving a two-layer slab.Comment: submitted to Math.Meth.Appl.Sc

The defect in the AT-like hamster cell mutants is complemented by mouse chromosome 9 but not by any of the human chromosomes

X-ray-sensitive Chinese hamster V79 cells mutants, V-C4, V-E5 and V-G8, show an abnormal response to X-ray-induced DNA damage. Like ataxia telangiectasia (AT) cells, they display increased cell killing, chromosomal instability and a diminished inhibition of DNA synthesis following ionizing radiation. To localize the defective hamster gene (XRCC8) on the human genome, human chromosomes were introduced into the AT-like hamster mutants, by microcell mediated chromosome transfer. Although, none of the human chromosomes corrected the defect in these mutants, the defect was corrected by a single mouse chromosome, derived from the A9 microcell donor cell line. In four independent X-ray-resistant microcell hybrid clones of V-E5, the presence of the mouse chromosome was determined by fluorescent in situ hybridization, using a mouse cot-1 probe. By PCR analysis with primers specific for different mouse chromosomes and Southern blot analysis with the mouse Ldlr probe, the mouse chromosome 9, was identified in all four X-ray-resistant hybrid clones. Segregation of the mouse chromosome 9 from these hamster-mouse microcell hybrids led to the loss of the regained X-ray-resistance, confirming that mouse chromosome 9 is responsible for complementation of the defect in V-E5 cells. The assignment of the mouse homolog of the ATM gene to mouse chromosome 9, and the presence of this mouse chromosome only in the radioresistant hamster cell hybrids suggest that the hamster AT-like mutants are homologous to AT, although they are not complemented by human chromosome 11

Understanding neurodevelopmental trajectories and behavioral profiles in SCN1A-related epilepsy syndromes

Background: A pathogenic variant in SCN1A can result in a spectrum of phenotypes, including Dravet syndrome (DS) and genetic epilepsy with febrile seizures plus (GEFS + ) syndrome. Dravet syndrome (DS) is associated with refractory seizures, developmental delay, intellectual disability (ID), motor impairment, and challenging behavior(1,2). GEFS + is a less severe phenotype in which cognition is often normal and seizures are less severe. Challenging behavior largely affects quality of life of patients and their families. This study describes the profile and course of the behavioral phenotype in patients with SCN1A-related epilepsy syndromes, explores correlations between behavioral difficulties and potential risk factors. Methods: Data were collected from questionnaires, medical records, and semi-structured interviews. Behavior difficulties were measured using the Adult/Child Behavior Checklist (C/ABCL) and Adult self-report (ASR). Other questionnaires included the Pediatric Quality of Life Inventory (PedsQL), the Functional Mobility Scale (FMS) and the Sleep Behavior Questionnaire by Simonds & Parraga (SQ-SP). To determine differences in behavioral difficulties longitudinally, paired T-tests were used. Pearson correlation and Spearman rank test were used in correlation analyses and multivariable regression analyses were employed to identify potential risk factors. Results: A cohort of 147 participants, including 107 participants with DS and 40 with genetic epilepsy with febrile seizures plus (GEFS + ), was evaluated. Forty-six DS participants (43.0 %) and three GEFS + participants (7.5 %) showed behavioral problems in the clinical range on the A/CBCL total problems scale. The behavioral profile in DS exists out of withdrawn behavior, aggressive behavior, and attention problems. In DS patients, sleep disturbances (β = 1.15, p < 0.001) and a lower age (β = -0.21, p = 0.001) were significantly associated with behavioral difficulties. Between 2015 and 2022, behavioral difficulties significantly decreased with age (t = -2.24, CI = -6.10 – −0.15, p = 0.04) in DS participants aging from adolescence into adulthood. A decrease in intellectual functioning (β = 3.37, p = 0.02) and using less antiseizure medications in 2022 than in 2015, (β = -1.96, p = 0.04), were identified as possible risk factors for developing (more) behavioral difficulties. Conclusions: These findings suggest that, in addition to epilepsy, behavioral difficulties are a core feature of the DS phenotype. Behavioral problems require personalized management and treatment strategies. Further research is needed to identify effective interventions

SOS: A Screening Instrument to Identify Children with Handwriting Impairments

Poor handwriting has been shown to be associated with developmental disorders such as Developmental Coordination Disorder, Attention Deficit Hyperactivity Disorder, autism, and learning disorders. Handwriting difficulties could lead to academic underachievement and poor self-esteem. Therapeutic intervention has been shown to be effective in treating children with poor handwriting, making early identification critical. The SOS test (Systematic Screening for Handwriting Difficulties) has been developed for this purpose. A child copies a sample of writing within 5 min. Handwriting quality is evaluated using six criteria and writing speed is measured. The Dutch SOS test was administered to 860 Flemish children (7-12 years). Inter-and intrarater reliability was excellent. Test-retest reliability was moderate. A correlation coefficient of 0.70 between SOS and "Concise Assessment Methods of Children Handwriting" test (Dutch version) confirmed convergent validity. The SOS allowed discrimination between typically developing children and children in special education, males and females, and different age groups