Sample handling of gastric tissue and O-glycan alterations in paired gastric cancer and non-tumorigenic tissues

Sample collection, handling and storage are the most critical steps for ensuring the highest preservation of specimens. Pre-analytical variability can influence the results as protein signatures alter rapidly after tissue excision or during long-term storage. Hence, we evaluated current state-of-the-art biobank preservation methods from a glycomics perspective and analyzed O-glycan alterations occurring in the gastric cancer tissues. Paired tumor and adjacent normal tissue samples were obtained from six patients undergoing gastric cancer surgery. Collected samples (n = 24) were either snap-frozen or heat stabilized and then homogenized. Glycans were released from extracted glycoproteins and analyzed by LC-MS/MS. In total, the relative abundance of 83 O-glycans and 17 derived structural features were used for comparison. There was no statistically significant difference found in variables between snap frozen and heat-stabilized samples, which indicated the two preservation methods were comparable. The data also showed significant changes between normal and cancerous tissue. In addition to a shift from high sialylation in the cancer area towards blood group ABO in the normal area, we also detected that the LacdiNAc epitope (N,N'-diacetyllactosamine) was significantly decreased in cancer samples. The O-glycan alterations that are presented here may provide predictive power for the detection and prognosis of gastric cancer

Proteomics of barley starch granules

Starches in various forms are an important part of the human diet. Furthermore, the use of starch as a renewable and biodegradable raw material for various industrial applications is becoming increasingly attractive. Non-food applications of starch make new demands on quality and type of starches produced. Ideally these new demands should be met by tailoring starches within the plant. A number of such in planta produced specialty starches exist today, e.g. high-amylose starch. Our aims with this Ph.D. project have been to increase the knowledge of starch synthesis and caryopsis development. We have attempted to do this primarily by investigating the proteins entrapped within the starch granule. This has been done using proteomic techniques. Our main focus has been on characterizing a barley mutant with high amylose starch, amo1. Apart from proteomic characterization of granule proteins we have also done comparative starch characterizations and assayed for starch branching activity in mutant and wild type seeds. We also investigated the occurrence of a caspase-like protease activity, a VEIDase, during caryopsis development. A protocol for extracting proteins from starch granules has been developed and a 2D proteomic reference map of integral granule proteins established. The molecular mechanism of a fragmentation of granule proteins induced during sample preparation has been found. An alternative sample preparation procedure has been established that reduces fragmentation considerably. A developmentally regulated caspase-like proteolytic activity, a VEIDase, with possible connection to developmental programmed cell death, has been characterized and found to be chiefly active during early stages of caryopsis development. The VEIDase activity has been localised to small vesicles in starchy endosperm cells

qPCR based mRNA quality score show intact mRNA after heat stabilization

Analysis of multiple analytes from biological samples can be challenging as different analytes require different preservation measures. Heat induced enzymatic inactivation is an efficient way to preserve proteins and their modifications in biological samples but RNA quality, as measured by RIN value, has been a concern in such samples. Here, we investigate the effect of heat stabilization compared with standard snap freezing on RNA quality using two RNA extraction protocols, QiaZol with and without urea pre-solubilization, and two RNA quality measurements: RIN value, as defined by the Agilent Bioanalyzer, and an alternative qPCR based method. DNA extraction from heat stabilized brain samples was also examined. The snap frozen samples had RIN values about 1 unit higher than heat stabilized samples for the direct QiaZol extraction but equal with stabilized samples using urea pre-solubilization. qPCR based RNA quality measurement showed no difference in quality between snap frozen and heat inactivated samples. The probable explanation for this discrepancy is that the RIN value is an indirect measure based on rRNA, while the qPCR score is based on actual measurement of mRNA quality. The DNA yield from heat stabilized brain tissue samples was significantly increased, compared to the snap frozen tissue, without any effects on purity or quality. Hence, heat stabilization of tissues opens up the possibility for a two step preservation protocol, where proteins and their modifications can be preserved in the first heat based step, while in a second step, using standard RNA preservation strategies, mRNA be preserved. This collection strategy will enable biobanking of samples where the ultimate analysis is not determined without loss of sample quality

GreenCharge : demotest i fält med elbuss

GreenCharge Sydost är en sammanslutning av regionförbund, kommuner, landsting och företag medett övergripande syfte att främja införandet av elfordon i främst sydöstra Sverige. Blekinge TekniskaHögskola är huvudman för projektet med ansvar för att driva forskningen, och Miljöfordon Syd attdriva den operativa projektledningen avseende demonstrationer och samverkan med intressenter.Under 2013 gjorde forskningen inom GreenCharge en beräkningsstudie som påvisade att elbussar urett livscykelperspektiv har mycket mindre utsläpp än dagens dieselbussar och totalkostnaden skullekunna bli 25 % lägre över en 8-års period i Karlskrona på linje 1 och 7, 21 % i Jönköping på linje 1och 3, samt 17 % i Sundsvall på linje 2 och 4. Detta gäller under antagande att bussen drivs med nygrön el, att realränteökningen blir 1 % per år samt att energi-pristrender sedan 10 år tillbaka fortsättergälla framöver.. Beräkningsstudien antog också efter simuleringar utifrån befintliga linjer och aktuellavärden från busstillverkare en energianvändning på 1,04 kWh/km för eldrift

A Novel WRKY Transcription Factor, SUSIBA2, Participates in Sugar Signaling in Barley by Binding to the Sugar-Responsive Elements of the iso1 Promoter

SURE (sugar responsive) is a cis element in plant sugar signaling. The SURE element was reported first for potato, in which it confers sugar responsiveness to the patatin promoter. A SURE binding transcription factor has not been isolated. We have isolated a transcription factor cDNA from barley and purified the corresponding protein. The transcription factor, SUSIBA2 (sugar signaling in barley), belongs to the WRKY proteins and was shown to bind to SURE and W-box elements but not to the SP8a element in the iso1 promoter. Nuclear localization of SUSIBA2 was demonstrated in a transient assay system with a SUSIBA2:green fluorescent protein fusion protein. Exploiting the novel transcription factor oligodeoxynucleotide decoy strategy with transformed barley endosperm provided experimental evidence for the importance of the SURE elements in iso1 transcription. Antibodies against SUSIBA2 were produced, and the expression pattern for susiba2 was determined at the RNA and protein levels. It was found that susiba2 is expressed in endosperm but not in leaves. Transcription of susiba2 is sugar inducible, and ectopic susiba2 expression was obtained in sugar-treated leaves. Likewise, binding to SURE elements was observed for nuclear extracts from sugar-treated but not from control barley leaves. The temporal expression of susiba2 in barley endosperm followed that of iso1 and endogenous sucrose levels, with a peak at ∼12 days after pollination. Our data indicate that SUSIBA2 binds to the SURE elements in the barley iso1 promoter as an activator. Furthermore, they show that SUSIBA2 is a regulatory transcription factor in starch synthesis and demonstrate the involvement of a WRKY protein in carbohydrate anabolism. Orthologs to SUSIBA2 were isolated from rice and wheat endosperm

GreenCharge : demotest i fält med elbuss

GreenCharge Sydost är en sammanslutning av regionförbund, kommuner, landsting och företag medett övergripande syfte att främja införandet av elfordon i främst sydöstra Sverige. Blekinge TekniskaHögskola är huvudman för projektet med ansvar för att driva forskningen, och Miljöfordon Syd attdriva den operativa projektledningen avseende demonstrationer och samverkan med intressenter.Under 2013 gjorde forskningen inom GreenCharge en beräkningsstudie som påvisade att elbussar urett livscykelperspektiv har mycket mindre utsläpp än dagens dieselbussar och totalkostnaden skullekunna bli 25 % lägre över en 8-års period i Karlskrona på linje 1 och 7, 21 % i Jönköping på linje 1och 3, samt 17 % i Sundsvall på linje 2 och 4. Detta gäller under antagande att bussen drivs med nygrön el, att realränteökningen blir 1 % per år samt att energi-pristrender sedan 10 år tillbaka fortsättergälla framöver.. Beräkningsstudien antog också efter simuleringar utifrån befintliga linjer och aktuellavärden från busstillverkare en energianvändning på 1,04 kWh/km för eldrift

Using collective expert judgements to evaluate quality measures of mass spectrometry images

International audienc