Bacillus thuringiensis is the most widely used bacterial bio-insecticide, and most insecticidal crystal protein-coding genes are located on plasmids. Most strains of B. thuringiensis harbor numerous diverse plasmids, although the plasmid copy numbers (PCNs) of all native plasmids in this host and the corresponding total plasmid DNA amount remains unknown. In this study, we determined the PCNs of 11 plasmids (ranging from 2 kb to 416 kb) in a sequenced B. thuringiensis subsp. kurstaki strain YBT-1520 using real-time qPCR. PCNs were found to range from 1.38 to 172, and were negatively correlated to plasmid size. The amount of total plasmid DNA (∼8.7 Mbp) was 1.62-fold greater than the amount of chromosomal DNA (∼5.4 Mbp) at the mid-exponential growth stage (OD600 = 2.0) of the organism. Furthermore, we selected three plasmids with different sizes and replication mechanisms to determine the PCNs over the entire life cycle. We found that the PCNs dynamically shifted at different stages, reaching their maximum during the mid-exponential growth or stationary phases and remaining stable and close to their minimum after the prespore formation stage. The PCN of pBMB2062, which is the smallest plasmid (2062 bp) and has the highest PCN of those tested, varied in strain YBT-1520, HD-1, and HD-136 (172, 115, and 94, respectively). These findings provide insight into both the total plasmid DNA amount of B. thuringiensis and the strong ability of the species to harbor plasmids.
A major safety accident will be triggered when the A-annular pressure value of high pressure, high productivity and high sulfur gas well exceeded the maximum allowable value. The A-annular pressure value of high pressure high productivity and high sulfur gas well once exceeded the allowable value will trigger a major safety accident. Therefore, this paper proposed a data mining based the gas well early warning strategy by analyzing the annular pressure mechanism and the change in the pattern of instantaneous gas volume, well temperature, and annular pressure in various conditions. To summarize, the law of gas well abnormal A-annular pressure is aimed at constructing a gas well safety warning rule for gas well stable production stage and shutdown period where the initial parameters setting in the early warning rule and adjustment optimization mechanism is also determined. Lastly, with the use of historical abnormal samples, the gas well early warning strategy bought up in this paper was verified. The example shown that compared with the traditional DCS system warning strategy, the gas well safety early warning strategy can identify the abnormal A-annular pressure phenomenon 82 h in advance thus achieving production safety management.
Bacillus thuringiensis serovar. finitimus strain YBT-020 is a typical strain with the spore-crystal association (SCA) phenotype. In our previous studies, plasmid curing experiment suggested that native plasmid pBMB28 of strain YBT-020 might contribute to the SCA phenotype. Thus, plasmid pBMB28 was cloned in order to isolate the genes related to SCA on pBMB28. Using shuttle vector pEMB0557, a shuttle genomic bacterial artificial chromosome (BAC) library of B. thuringiensis strain YBT-020 was constructed. The plasmid pBMB231 containing crystal protein gene cry28Aa, which was located on plasmid pBMB28, was screened out. By SDS-PAGE analysis and microscopic observation, we discovered the recombinant strain BMB231 that originated from the electrotransfer strain BMB171 with pBMB231 could produce Cry28Aa protein. With the chromosome walking strategy and terminal sequencing of pBMB231, four clones covering the full length of plasmid pBMB28 were screened out from this BAC library. With pulsed gel analysis of the four BAC clones and terminal sequencing, the size of the plasmid was calculated to be 140 kb. This study additionally revealed that we could clone a large plasmid from B. thuringiensis by genomic BAC library construction and overlaping fragment screening.
The proposed paper is going to address the development of single point mooring FPSO (Floating Production, Storage and Offloading) monitoring and forecast system design. With 17 FPSOs deployed in both Bohai Bay and South China Sea, CNOOC owns one of the largest FPSO fleet in the world. Most of those FPSOs have been or will be moored to the seabed for decades. The extreme response during storm conditions could cause serious environmental problem, asset loss, personnel safety etc. In order to timely understand the tanker operation conditions and avoid potential risk of system failure when experiencing hurricanes, a monitoring and forecast system is developed for FPSO to monitor the environment conditions, tanker motions, green water, mooring tensions, FPSO heading and to predict the extreme mooring tensions and global motions before typhoon coming. The forecast system could further suggest the optimum loading condition for minimizing the extreme mooring tension and tanker motions to enhance the safe operation. In this paper, we take the Internal Turret Mooring FPSO 111 and the Submerged Soft Yoke Mooring FPSO 112 as the examples to introduce the design technology of the system. Through the integrated onboard interface information, the personnel could proactively take actions to mitigate the tensions on mooring lines and vessel motions. Furthermore, the measured mooring line tension, motion and environment history could assist the numerical studies of global performance. The details of designing or selecting the measuring and monitoring equipment, theory background of forecast system and the integrated onboard interface will be described.
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