Detecting imbalanced expression of SNP alleles by minisequencing on microarrays

BACKGROUND: Each of the human genes or transcriptional units is likely to contain single nucleotide polymorphisms that may give rise to sequence variation between individuals and tissues on the level of RNA. Based on recent studies, differential expression of the two alleles of heterozygous coding single nucleotide polymorphisms (SNPs) may be frequent for human genes. Methods with high accuracy to be used in a high throughput setting are needed for systematic surveys of expressed sequence variation. In this study we evaluated two formats of multiplexed, microarray based minisequencing for quantitative detection of imbalanced expression of SNP alleles. We used a panel of ten SNPs located in five genes known to be expressed in two endothelial cell lines as our model system. RESULTS: The accuracy and sensitivity of quantitative detection of allelic imbalance was assessed for each SNP by constructing regression lines using a dilution series of mixed samples from individuals of different genotype. Accurate quantification of SNP alleles by both assay formats was evidenced for by R(2 )values > 0.95 for the majority of the regression lines. According to a two sample t-test, we were able to distinguish 1–9% of a minority SNP allele from a homozygous genotype, with larger variation between SNPs than between assay formats. Six of the SNPs, heterozygous in either of the two cell lines, were genotyped in RNA extracted from the endothelial cells. The coefficient of variation between the fluorescent signals from five parallel reactions was similar for cDNA and genomic DNA. The fluorescence signal intensity ratios measured in the cDNA samples were compared to those in genomic DNA to determine the relative expression levels of the two alleles of each SNP. Four of the six SNPs tested displayed a higher than 1.4-fold difference in allelic ratios between cDNA and genomic DNA. The results were verified by allele-specific oligonucleotide hybridisation and minisequencing in a microtiter plate format. CONCLUSIONS: We conclude that microarray based minisequencing is an accurate and accessible tool for multiplexed screening for imbalanced allelic expression in multiple samples and tissues in parallel

Welcome to the party!: Unpacking Party Organizations

First paragraph: As an organizational species, political parties seem to face impending extinction. No matter what yardstick we use to measure their vitality, political parties currently display an undeniable image of terminal crisis. Party membership is approaching rock bottom in most corners of the world, particularly in countries like France and the UK where less than two percent of the population are registered as rank and file (van Biezen et al., 2012). Similarly, voter turnout has plummeted worldwide since the middle of the twentieth century, currently reaching a level well below 70 percent (Solijonov, 2016). Voters' tendency to identify with specific parties is likewise declining due to the reconfiguration of class-consciousness and the emergence of more ‘liquid loyalties’ in the electorate (Ignazi, 2017: 201). Finally, people’s trust in political parties is at an all-time low, with politicians deemed less trustworthy than complete strangers and more dishonest than second-hand car dealers (Newton et al., 2017). As such, it seems fair to conclude, as many have recently done, that the party is over (e.g. Holloway, 2002; Day, 2005; Rosanvallon, 2008; Castells, 2012; della Porta, 2013; Tormey, 2015; Hardt and Negri, 2017)

Political parties and Organization Studies: The party as a critical case of organizing

Organization scholars have extensively studied both the politics of organization and the organization of politics. Contributing to the latter, we argue for further and deeper consideration of political parties, since: (1) parties illuminate organizational dynamics of in- and exclusion; (2) internal struggles related to the constitution of identities, practices, and procedures are accentuated in parties; (3) the study of parties allow for the isolation of processes of normative and affective commitment; (4) parties prioritize and intensify normative control mechanisms; (5) party organizing currently represents an example of profound institutional change, as new (digital) formations challenge old bureaucratic models. Consequently, we argue that political parties should be seen as ‘critical cases’ of organizing, meaning that otherwise commonplace phenomena are intensified and exposed in parties. This allows researchers to use parties as magnifying glasses for zooming-in on organizational dynamics that may be suppressed or concealed by the seemingly non-political façade of many contemporary organizations. In conclusion, we argue that organization scholars are in a privileged position to investigate how political parties function today and how their democratic potential can be improved in the future. To this end, we call on Organization and Management Studies to engage actively with alternative parties in an attempt to explore and promote progressive change within the formal political system.Output Status: Forthcoming/Available Onlin

Welcome to the party!: Unpacking Party Organizations

First paragraph: As an organizational species, political parties seem to face impending extinction. No matter what yardstick we use to measure their vitality, political parties currently display an undeniable image of terminal crisis. Party membership is approaching rock bottom in most corners of the world, particularly in countries like France and the UK where less than two percent of the population are registered as rank and file (van Biezen et al., 2012). Similarly, voter turnout has plummeted worldwide since the middle of the twentieth century, currently reaching a level well below 70 percent (Solijonov, 2016). Voters' tendency to identify with specific parties is likewise declining due to the reconfiguration of class-consciousness and the emergence of more ‘liquid loyalties’ in the electorate (Ignazi, 2017: 201). Finally, people’s trust in political parties is at an all-time low, with politicians deemed less trustworthy than complete strangers and more dishonest than second-hand car dealers (Newton et al., 2017). As such, it seems fair to conclude, as many have recently done, that the party is over (e.g. Holloway, 2002; Day, 2005; Rosanvallon, 2008; Castells, 2012; della Porta, 2013; Tormey, 2015; Hardt and Negri, 2017)

Carriage of methicillin-resistant Staphylococcus pseudintermedius in dogs--a longitudinal study

<p>Abstract</p> <p>Background</p> <p>Methicillin-resistant <it>S. pseudintermedius </it>strains (MRSP) are reported with increasing frequency in bacterial cultures from dogs. The objectives of this study were to determine whether MRSP could be found in dogs several months after a clinically apparent infection and whether the length of carriage varied depending on systemic antimicrobial treatment, diagnosis at time of the first positive MRSP culture and the presence of skin disease or wounds. Thirty-one dogs previously diagnosed with a clinical infection were sampled repeatedly for a minimum of eight months or, with the exception of two dogs, until two consecutive negative results were obtained. Five specified locations were sampled, and the results were evaluated to determine future recommendations concerning sample strategies when screening for MRSP carriage. Information was collected from medical records and questionnaires to evaluate factors that may influence length of carriage.</p> <p>Results</p> <p>The overall median length of MRSP carriage was 11 months (48 weeks). The presence of wounds and signs of dermatitis did not influence length of carriage. Systemic treatment for three weeks or longer with antimicrobial agents to which the bacterium was resistant was associated with prolonged carriage compared to dogs treated for a shorter period of time. Three of five dogs treated with an antimicrobial to which their MRSP-isolates were susceptible (tetracycline) were found to still be MRSP-positive when sampled after the end of treatment. Wound samples had the highest positive MRSP yield (81%) for the positive sample sites, compared to less than 70% for each of the other four sample sites. Cultures from the nostrils were less likely to detect MRSP carriage relative to the pharynx, perineum, wounds and the corner of the mouth.</p> <p>Conclusions</p> <p>Dogs can carry MRSP for more than a year after a clinically apparent infection. Systemic antimicrobial treatment of infections with antimicrobial agents to which the MRSP-bacteria are resistant should be avoided when possible in dogs with possible or confirmed MRSP carriage or infection, since it may prolong time of MRSP carriage. Simultaneous sampling of pharynx, perineum, and the corner of the mouth as well as wounds when present is recommended when screening for MRSP. Cultures from nostrils were shown to be less likely to detect MRSP carriage.</p

Мотивация и ее методы социально-психологического стимулирования

Материалы XVIII Междунар. науч.-техн. конф. студентов, аспирантов и молодых ученых, Гомель, 26–27 апр. 2018 г

Using Minisequencing Technology for Analysing Genetic Variation in DNA and RNA

In this thesis, the four-color fluorescence tag-microarray minisequencing system pioneered by our group was further developed and applied for analysing genetic variation in human DNA and RNA. A SNP marker panel representing different chromosomal regions was established and used for identification of informative SNP markers for monitoring chimerism after stem cell transplantation (SCT). The success of SCT was monitored by measuring the allelic ratios of informative SNPs in follow-up samples from nine patients with leukaemia. The results agreed with data obtained using microsatellite markers. Further the same SNP marker panel was used for evaluation of two whole genome amplification methods, primer extension preamplification (PEP) and multiple displacement amplification (MDA) in comparison with genomic DNA with respect to SNP genotyping success and accuracy in tag-array minisequencing. Identical results were obtained from MDA products and genomic DNA. The tag-microarray minisequencing system was also established for multiplexed quantification of imbalanced expression of SNP alleles. Two endothelial cell lines and a panel of ten coding SNPs in five genes were used as model system. Six heterozygous SNPs were genotyped in RNA (cDNA) from the cell lines. Comparison of the relative amounts of the SNPs alleles in cDNA to heterozygote SNPs in genomic DNA displayed four SNPs with significant imbalanced expression between the SNP alleles. Finally, the tag-array minisequencing system was modified for detection of splice variants in mRNA from five leukaemia cell lines. A panel of 20 cancer-related genes with 74 alternatively splice variants was screened. Over half of the splice variants were detected in the cell lines, and similar alternative splicing patterns were observed in each cell line. The results were verified by size analysis of the PCR product subjected to the minisequencing primer extension reaction. The data from both methods agreed well, evidencing for a high sensitivity of our system