Badania nad strukturą i mikroflorą ziarn kefirowych w mikroskopie elektronowym

It was found out that participation of lactobacilli, streptococci and yeasts in kefir grailns' microflora amounted to 66, 16 and 18% respectively. Scanning and transmission microscopy revealed that lactic acid bacteria formed microcolonies mainly inside the grains and yeasts at the surface layers.W pracy określono proporcje pomiędzy różnymi formami morfologicznymi drobnoustrojów występujących w ziarnach kefirowych oraz badano strukturę i budowę ziarn w mikroskopie elektronowym. Przy liczeniu bakterii jako jednostkę traktowano zarówno komórki pojedyncze, jak też dwoinki i łańcuszki, przy liczeniu drożdży - poszczególne komórki. Liczenie komórek w preparatach barwionych sporządzonych z ziarn kefirowych rozpuszczonych w O, 1 N NaOH wykazało, że liczbowo pałeczki stanowiły średnio 66% mikroflory (wahania 62-69%), zierniaki 16%(wahania 11-22%), a drożdże 18% (wahania 16-20%). Obserwacje przeprowadzone w elektronowym mikroskopie skaningowym pozwoliły stwierdzić występowanie na powierzchni ziarn dużych skupień komórek drożdży i pałeczek,. co sugeruje, że rozwijają się one w mikrokoloniach tworzących masę ziarna. Ziarniaki występowały w obrębie tych skupień jako dwoinki i krótkie łańcuszki (rys. 2). W barwionych octanem uranylu ultracienkich skrawkach z ziarn kefirowych obserwowanych w elektronowym mikroskopie transmisyjnym zaobserwowano wewnątrz ziarn głównie komórki bakterii. Drożdże występowały nielicznie, co wskazuje na fakt silniejszego ich rozwoju w powierzchniowych częściach ziarn. Komórki drobnoustrojów zatopione były w bezpostaciowej substancji (rys. 3 i 4)

Urea supplementation improves mRNA in vitro transcription by decreasing both shorter and longer RNA byproducts

ABSTRACTThe current letter to the editor describes the presence of RNA byproducts in small-scale in vitro transcription (IVT) reactions as evaluated by capillary gel electrophoresis, asymmetric flow field flow fractionation, immunoblotting, cell-free translation assays, and in IFN reporter cells. We compare standard T7 RNA polymerase (RNAP) based IVT reactions to two recently described protocols employing either urea supplementation or using the VSW3 RNAP. Our results indicate that urea supplementation yields considerably less RNA byproducts and positively affects the overall number of full-length transcripts. In contrast, VSW3 IVT reactions demonstrated a low yield and generated a higher fraction of truncated transcripts. Lastly, both urea mRNA and VSW3 mRNA elicited considerably less IFN responses after transfection in mouse macrophages

Integrator is a genome-wide attenuator of non-productive transcription

Termination of RNA polymerase II (RNAPII) transcription in metazoans relies largely on the cleavage and polyadenylation (CPA) and integrator (INT) complexes originally found to act at the ends of protein-coding and small nuclear RNA (snRNA) genes, respectively. Here, we monitor CPA- and INT-dependent termination activities genome-wide, including at thousands of previously unannotated transcription units (TUs), producing unstable RNA. We verify the global activity of CPA occurring at pA sites indiscriminately of their positioning relative to the TU promoter. We also identify a global activity of INT, which is largely sequence-independent and restricted to a ~3-kb promoter-proximal region. Our analyses suggest two functions of genome-wide INT activity: it dampens transcriptional output from weak promoters, and it provides quality control of RNAPII complexes that are unfavorably configured for transcriptional elongation. We suggest that the function of INT in stable snRNA production is an exception from its general cellular role, the attenuation of non-productive transcription

Urea supplementation improves mRNA in vitro transcription by decreasing both shorter and longer RNA byproducts

The current letter to the editor describes the presence of RNA byproducts in small-scale in vitro transcription (IVT) reactions as evaluated by capillary gel electrophoresis, asymmetric flow field flow fractionation, immunoblotting, cell-free translation assays, and in IFN reporter cells. We compare standard T7 RNA polymerase (RNAP) based IVT reactions to two recently described protocols employing either urea supplementation or using the VSW3 RNAP. Our results indicate that urea supplementation yields considerably less RNA byproducts and positively affects the overall number of full-length transcripts. In contrast, VSW3 IVT reactions demonstrated a low yield and generated a higher fraction of truncated transcripts. Lastly, both urea mRNA and VSW3 mRNA elicited considerably less IFN responses after transfection in mouse macrophages.</p

Cryo-XPS for Surface Characterization of Nanomedicines

Nanoparticles used for medical applications commonly possess coatings or surface functionalities intended to provide specific behavior in vivo, for example, the use of PEG to provide stealth properties. Direct, quantitative measurement of the surface chemistry and composition of such systems in a hydrated environment has thus far not been demonstrated, yet such measurements are of great importance for the development of nanomedicine systems. Here we demonstrate the first use of cryo-XPS for the measurement of two PEG-functionalized nanomedicines: a polymeric drug delivery system and a lipid nanoparticle mRNA carrier. The observed differences between cryo-XPS and standard XPS measurements indicate the potential of cryo-XPS for providing quantitative measurements of such nanoparticle systems in hydrated conditions