To establish a constitutive, high-efficiency expression system for Bacillus pumilus (B.P), we cloned random chromosomal DNA into promoter probe shuttle vector ECE7 and selected for strong promoter activity by chloramphenicol resistance of transformed B. pumilus cells. The nucleotide sequences of nine chromosomal fragments were determined. These DNA fragments range from 300 to 2200 bp in size. The transcription strength of these promoters was estimated by determination of CAT enzyme production in both E. coli and B. pumilus. Transcription start (TS) sites of the cat mRNA were located by primer extension by using total RNA. Preliminary analysis showed that three of the promoter sequences contain -35 and -10 regions like E. coli RNA polymerase sigma70 and B. subtilis sigma43 consensus sequences. One is similar to B. subtilis sigma29, the other two have no conserved sequences like any of the typical consensus sequences of the known sigma factors so far. To estimate the feasibility of the utilization of these promoters, one promoter fragment was subcloned and used to drive the expression of green fluorescent protein (GFP) in B. pumilus cells. This is the first report of B. pumilus promoters randomly cloning from total DNA and molecular analysis of their consensus sequences.

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The transport properties of chloride ions in cement-based materials are one of the major deterioration mechanisms for reinforced concrete (RC) structures. This paper investigates the influence of pore size and fatigue loading on the transport properties of NaCl in C-S-H nanopores using molecular dynamics (MD) simulations. Molecular models of C-S-H, NaCl solution, and C-S-H nanopores with different pore diameters are established on a microscopic scale. The distribution of the chloride ion diffusion rate and the diffusion coefficient of each particle are obtained by statistically calculating the variation of atomic displacement with time. The results indicate that the chloride ion diffusion rate perpendicular to C-S-H nanopores under fatigue loading is 4 times faster than that without fatigue loading. Moreover, the diffusion coefficient of water molecules and chloride ions in C-S-H nanopores increases under fatigue loading compared with those without fatigue loading. The diffusion coefficient of water molecules in C-S-H nanopores with a pore size of 3 nm obtained from the MD simulation is 1.794 × 10−9 m2/s, which is slightly lower than that obtained from the experiment.

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