Multiplex gene removal by two-step polymerase chain reactions

Precise DNA manipulation is critical for molecular biotechnology. Restriction enzyme-based approaches are limited by their requirement of specific enzyme sites. Restriction-free cloning has greatly improved the flexibility and speed of precise DNA assembly. Most of these approaches focus on DNA assembly rather than gene removal. Here we present a polymerase chain reaction (PCR)-based cloning method that allows removal of multiple gene segments from plasmids without using restriction enzymes and thermostable ligase. We demonstrate simultaneous removal of three gene segments from a plasmid. This approach could be beneficial to DNA library construction, genetic and protein engineering, and synthetic biology.     

CO2 Saturation and Trophic Shift Induced by Microbial Metabolic Processes in a River-Dominated Ocean Margin (Tropical Shallow Lagoon,Chilika, India)

An effort has been made for the first time in Asia's largest brackish water lagoon, Chilika, to investigate the spatio-temporal variability in primary productivity (PP), bacterial productivity (BP), bacterial abundance (BA), bacterial respiration (BR) and bacterial growth efficiency (BGE) in relation to partial pressure of CO₂ (pCO₂) and CO₂ air–water flux and the resultant trophic switchover. Annually, PP ranged between 24 and 376 µg C L^?1 d^?1 with significantly low values throughout the monsoon (MN), caused by light limitation due to inputs of riverine suspended matter. On the contrary, BP and BR ranged from 11.5 to 186.3 µg C L^?1 d^?1 and from 14.1 to 389.4 µg C L^?1 d^?1, respectively, with exceptionally higher values during MN. A wide spatial and temporal variation in the lagoon trophic status was apparent from BP/PP (0.05–6.4) and PP/BR (0.10–18.2) ratios. The seasonal shift in net pelagic production from autotrophy to heterotrophy due to terrestrial organic matter inputs via rivers, enhanced the bacterial metabolism during the MN, as evident from the high pCO₂ (10,134 µatm) and CO₂ air–water flux (714 mm m^?2 d^?1). Large variability in BGE and BP/PP ratios especially during MN led to high bacteria-mediated carbon fluxes which was evident from significantly high bacterial carbon demand (BCD >100% of PP) during this season. This suggested that the net amount of organic carbon (either dissolved or particulate form) synthesized by primary producers in the lagoon was not sufficient to satisfy the bacterial carbon requirements. Lagoon sustained low to moderate autotrophic–heterotrophic coupling with annual mean BCD of 231% relative to the primary production, which depicted that bacterioplankton are the mainstay of the lagoon biogeochemical cycles and principal players that bring changes in trophic status. Study disclosed that the high CO₂ supersaturation and oxygen undersaturation during MN was attributed to the increased heterotrophic respiration (in excess of PP) fuelled by allochthonous organic matter. On a spatial scale, lagoon sectors such as south sector, central sector and outer channel recorded “net autotrophic,” while the northern sector showed “net heterotrophic” throughout the study period.     

Metal Chelates as Fungicides

A remarkable improvement in the emulsifying properties of proteins was observed in mixtures of serum albumin-dextran sulfate, lysozyme-dextran sulfate, serum albumin-chondroitin sulfate, α-lactalbumin-chondroitin sulfate and lysozyme-chondroitin sulfate. The emulsifying properties of these protein-polysaccharide complexes were affected by the molecular size of the polysaccharide and by such conditions as the presence of salt, acidic or alkaline pH, and heating. On the other hand, the emulsifying properties of ovalbumin-dextran hybrids prepared by covalent attachment were more enhanced in alkaline pH or by heating. The molecular size of the ovalbumin-dextran hybrids greatly affected the emulsifying properties.     

Picturing beach erosion and deposition trends using PSInSAR: an example from the non-barred southern west coast of India

Coastal dynamics can rapidly alter beach morphology. In some places, such as the non-barred southern west coast of India, studying changes to beach morphology is a relatively arduous task. Persistent Scatterer Interferometric Synthetic Aperture Radar (PSInSAR), a remote sensing technique that utilizes stacks of radar images for accurate long-term monitoring of ground features, allows for detailed observations of coastal morphodynamics. Twenty-two single look complex (SLC) synthetic aperture radar (SAR) Sentinel-1A images, acquired from 4 March 2016 to 3 June 2017, are used to monitor geomorphological processes such as coastal erosion and deposition. Ground deformation measurements from PSInSAR processing shows the coast stretching?~?70 km between Thaickal and Munambam is highly dynamic, characterized by phases of erosion and deposition. The highest negative displacement of ? 24.9 mm at Thaickal versus the?+?7.6 mm at Chellanam in the north show the co-existing milieus of erosion and deposition. PSInSAR results concur with corresponding Google Earth images. In addition, beach sediment texture and scanning electron microscope grain micro-texture in the beach segment further corroborate temporal phases of erosion and deposition. Two locations are identified as typical erosional sites, while one location typified deposition. Erosion and deposition or rebuilding of beaches, usually correspond respectively with the onset and offset of SW monsoon wave climate in the Arabian Sea. When zones of deposition are mainly located in the proximity of river and/or lake inlets, stretches characterized by erosion are distal to inlets. Linear regression analyses of displacement–time series plots were used to identify general erosion or depositional regimes along beach segments. Results from this study illustrate how PSInSAR is a capable and reliable processing tool for monitoring temporal phases of coastal morpho-dynamics.

     

Bringing Conducting Polymers to High Order: Toward Conductivities beyond 105 S cm−1 and Thermoelectric Power Factors of 2 mW m−1 K−2

Here, an effective design strategy of polymer thermoelectric materials based on structural control in doped polymer semiconductors is presented. The strategy is illustrated for two archetypical polythiophenes, e.g., poly(2,5‐bis(3‐dodecyl‐2‐thienyl)thieno[3,2‐b]thiophene) (C₁₂‐PBTTT) and regioregular poly(3‐hexylthiophene) (P3HT). FeCl₃ doping of aligned films results in charge conductivities up to 2 × 10⁵ S cm^?1 and metallic‐like thermopowers similar to iodine‐doped polyacetylene. The films are almost optically transparent and show strongly polarized near‐infrared polaronic bands (dichroic ratio >10). The comparative study of structure–property correlations in P3HT and C₁₂‐PBTTT identifies three conditions to obtain conductivities beyond 10⁵ S cm^?1: i) achieve high in‐plane orientation of conjugated polymers with high persistence length; ii) ensure uniform chain oxidation of the polymer backbones by regular intercalation of dopant molecules in the polymer structure without disrupting alignment of π‐stacked layers; and iii) maintain a percolating nanomorphology along the chain direction. The highly anisotropic conducting polymer films are ideal model systems to investigate the correlations between thermopower S and charge conductivity σ. A scaling law S ∝ σ^?1/4 prevails along the chain direction, but a different S ∝ ?ln(σ) relation is observed perpendicular to the chains, suggesting different charge transport mechanisms. The simultaneous increase of charge conductivity and thermopower along the chain direction results in a substantial improvement of thermoelectric power factors up to 2 mW m^?1 K^?2 in C₁₂‐PBTTT.