RT-Bst: an integrated approach for reverse transcription and enrichment of cDNA from viral RNA

The synthesis of cDNA from RNA is challenging due to the inefficiency of reverse transcription (RT). In order to address this, a method was developed known as RT-Bst for sequential RT of RNA and Bst DNA polymerase amplification for enrichment of cDNA in a single tube reaction. Using genomic RNA from bacteriophage MS2, the yield of cDNA produced by RT alone and RT-Bst were compared by analysis of PCR-amplified products. Using random primers a superior performance was observed when amplifying MS2 RNA following RT-Bst compared to RT alone, indicating that greater quantities of cDNA were present after RT-Bst. RT-Bst was also compared with RT alone for their relative ability to produce sufficient cDNA to amplify 8 target regions spanning the respiratory syncytial virus (RSV) genome. Six out of 8 targets were amplified consistently by PCR subsequent to RT-Bst amplification whereas only 3 out of 8 targets could be amplified after RT alone. RSV sequences were selectively amplified using RSV specific primers from a mixed template containing an excess of MS2 RNA in a RT-Bst reaction without amplifying MS2 sequences. This suggests that RT-Bst can be used to amplify RNA sequences non-specifically using random primers and specifically using sequence specific primers and enhances the yield of cDNA when compared to RT alone.

Strength improvement of adhesively-bonded joints using a reverse-bent geometry

Adhesive bonding of components has become more efficient in recent years due to the developments in adhesive technology, which has resulted in higher peel and shear strengths, and also in allowable ductility up to failure. As a result, fastening and riveting methods are being progressively replaced by adhesive bonding, allowing a big step towards stronger and lighter unions. However, single-lap bonded joints still generate substantial peel and shear stress concentrations at the overlap edges that can be harmful to the structure, especially when using brittle adhesives that do not allow plasticization in these regions. In this work, a numerical and experimental study is performed to evaluate the feasibility of bending the adherends at the ends of the overlap for the strength improvement of single-lap aluminium joints bonded with a brittle and a ductile adhesive. Different combinations of joint eccentricity were tested, including absence of eccentricity, allowing the optimization of the joint. A Finite Element stress and failure analysis in ABAQUS® was also carried out to provide a better understanding of the bent configuration. Results showed a major advantage of using the proposed modification for the brittle adhesive, but the joints with the ductile adhesive were not much affected by the bending technique.

Reverse problem-based learning - a case study with a Braille machine

Engineering Education includes not only teaching theoretical fundamental concepts but also its verification during practical lessons in laboratories. The usual strategies to carry out this action are frequently based on Problem Based Learning, starting from a given state and proceeding forward to a target state. The possibility or the effectiveness of this procedure depends on previous states and if the present state was caused or resulted from earlier ones. This often happens in engineering education when the achieved results do not match the desired ones, e.g. when programming code is being developed or when the cause of the wrong behavior of an electronic circuit is being identified. It is thus important to also prepare students to proceed in the reverse way, i.e. given a start state generate the explanation or even the principles that underlie it. Later on, this sort of skills will be important. For instance, to a doctor making a patient?s story or to an engineer discovering the source of a malfunction. This learning methodology presents pedagogical advantages besides the enhanced preparation of students to their future work. The work presented on his document describes an automation project developed by a group of students in an engineering polytechnic school laboratory. The main objective was to improve the performance of a Braille machine. However, in a scenario of Reverse Problem-Based learning, students had first to discover and characterize the entire machine's function before being allowed (and being able) to propose a solution for the existing problem.

Time to Left Ventricular Reverse Remodeling after Cardiac Resynchronization Therapy: Better Late than Never

INTRODUCTION: Left ventricular reverse remodeling (LVRR), defined as reduction of end-diastolic and end-systolic dimensions and improvement of ejection fraction, is associated with the prognostic implications of cardiac resynchronization therapy (CRT). The time course of LVRR remains poorly characterized. Nevertheless, it has been suggested that it occurs ≤6 months after CRT. OBJECTIVE: To characterize the long-term echocardiographic and clinical evolution of patients with LVRR occurring >6 months after CRT and to identify predictors of a delayed LVRR response. METHODS: A total of 127 consecutive patients after successful CRT implantation were divided into three groups according to LVRR response: Group A, 19 patients (15%) with LVRR after >6 months (late LVRR); Group B, 58 patients (46%) with LVRR before 6 months (early LVRR); and Group C, 50 patients (39%) without LVRR during follow-up (no LVRR). RESULTS: The late LVRR group was older, more often had ischemic etiology and fewer patients were in NYHA class ≤II. Overall, group A presented LVRR between group B and C. This was also the case with the percentage of clinical response (68.4% vs. 94.8% vs. 38.3%, respectively, p<0.001), and hospital readmissions due to decompensated heart failure (31.6% vs. 12.1% vs. 57.1%, respectively, p<0.001). Ischemic etiology (OR 0.044; p=0.013) and NYHA functional class

Digital interconnectivity as a driver for port and logistics competitiveness

Creating an innovative tool that takes advantage of digital interconnectivity between shipping agencies and husbandry services suppliers was the starting point. But the main purpose of this paper is to figure out if that represents a business opportunity. It describes the preliminary stages undertaken, as the connections with the main potential providers of the husbandry services. This was carried out as a qualitative research, based on interviews given by shipping agencies that contributed as a source of data about their activities but also to survey their acceptance of the concept that could change the way of doing business in this area.At the same time, inquiries have been made to build financial scenarios that show the costs and revenue streams allocated to this project. Considering the data collected from the main players in husbandry services and the different outcomes, the feasibility of this project is assessed. Even though the paradigm was well received by all the firms contacted, the development costs turn out to be the main threat to the project so further steps are advised.

Market Opportunities Analysis for Logistics Solutions Provider Case Danzas Solutions

Tutkielman tavoitteena oli suunitella markkina-analyysi ja sitä käyttäen määritellä Danzasin Solutions liiketoimintayksikön mahdollisuuksia laajentaa toimintaansa Suomen terveydenhuolto/ lääke- ja elektroniikka/ telekommunikaatiosektoreilla. Danzas Solutions toimisi logistiikkaintegraattorina toimittamalla kokonaisvaltaisia logistisia ratkaisuja. Tutkimus tehtiin haastattelemalla neljää Danzasin asiakasta. Mahdollisimman laajan ja yleistettävän tuloksen saamiseksi haastateltavat valitiin logistiikan ulkoistamisasteeltaan vaihteleviksi. Tutkimus tehtiin ensin tutkimalla sekundääristä informaatiota markkinoilta ja täydentämällä sitä haastatteluista saadulla kvalitatiivisella primääri informaatiolla ja case yrityksen sisäisellä analyysillä. Painopiste tutkimuksessa oli selvittää yritysten ulkoistamiseen ja logistiikkakumppanin valintaan vaikuttavia kriteereitä. Logistiikkaintegraattorille suurimmat mahdollisuudet löytyvät elektroniikka/ telekommunikaatiosektorilta. Logistiikka tällä sektorilla on vaativaa ja yrityksillä ei ole resursseja hallita sitä itse. Terveydenhuolto/ lääkesektorilla yrityksillä on yleensä muut keinot jakeluketjun hallintaan ja ne eivät tarvitse logistiikkaintegraattorin palveluita.

The Albion, or British, Colonial and Foreign Weekly Gazette Vol. 5, No. 18- May 2, 1846. Article on Major General Isaac Brock and the 41st Regiment on the second (reverse) page.

A weekly paper published from 1822 to 1856.

DNA-Directed DNA Polymerases Evolve From Reverse Transcriptase

Scientists have been debating for decades the origin of life on earth. A number of hypotheses were proposed as to what emerged first RNA or DNA; with most scientists are in favour of the "RNA World" hypothesis. Assuming RNA emerged first, it fellow that the RNA polymerases would've appeared before DNA polymerases. Using recombinant DNA technology and bioinformatics we undertook this study to explore the relationship between RNA polymerases, reverse transcriptase and DNA polymerases. The working hypothesis is that DNA polymerases evolved from reverse transcriptase and the latter evolved from RNA polymerases. If this hypothesis is correct then one would expect to find various ancient DNA polymerases with varying level of reverse transcriptase activity. In the first phase of this research project multiple sequence alignments were made on the protein sequence of 32 prokaryotic DNA-directed DNA polymerases originating from 11 prokaryotic families against 3 viral reverse transcriptase. The data from such alignments was not very conclusive. DNA polymerases with higher level of reverse transcriptase activity were non-confined to ancient organisms, as one would've expected. The second phase of this project was focused on conditions that may alter the DNA polymerase activity. Various reaction conditions, such as temperature, using various ions (Ni2+, Mn2+, Mg2+) were tested. Interestingly, it was found that the DNA polymerase from the Thermos aquatics family can be made to copy RNA into DNA (i.e. reverse transcriptase activity). Thus it was shown that under appropriate conditions (ions and reactions temperatures) reverse transcriptase activity can be induced in DNA polymerase. In the third phase of this study recombinant DNA technology was used to generate a chimeric DNA polymerase; in attempts to identify the region(s) of the polymerase responsible for RNA-directed DNA polymerase activity. The two DNA polymerases employed were the Thermus aquatic us and Thermus thermophiles. As in the second phase various reaction conditions were investigated. Data indicated that the newly engineered chimeric DNA polymerase can be induced to copy RNA into DNA. Thus the intrinsic reverse transcriptase activity found in ancient DNA polymerases was localized into a domain and can be induced via appropriate reaction conditions.