Fusion of SpCas9 to E.coli Rec A protein enhances CRISPR-Cas9 mediated gene knockout in mammalian cells

Mammalian cells repair double-strand DNA breaks (DSB) by a range of different pathways following DSB induction by the engineered clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein Cas9. While CRISPR-Cas9 thus enables predesigned modifications of the genome, applications of CRISPR-Cas9-mediated genome-editing are frequently hampered by the unpredictable and varying pathways for DSB repair in mammalian cells. Here we present a strategy of fusing Cas9 to recombinant proteins for fine-tuning of the DSB repair preferences in mammalian cells. By fusing Streptococcus Pyogenes Cas9 (SpCas9) to the recombinant protein A (Rec A, NP_417179.1) from E. coli, we create a recombinant Cas9 protein (rSpCas9) which enhances the generation of indel mutations at DSB sites in mammalian cells, increases the frequency of DSB repair by homology-directed single-strand annealing (SSA), and represses homology-directed gene conversion by approximately 33%. Our study thus proves for the first time that fusing SpCas9 to recombinant proteins can influence the balance between DSB repair pathways in mammalian cells. This approach may form the basis for further investigations of the applications of recombinant Cas9 proteins to fine-tuning DSB repair pathways in eukaryotic cells

Fish assemblages in the mangrove creeks of northern and southern Taiwan

A bimonthly study of the spatial variations in fish assemblages in the six mangrove creeks along the western coast of Taiwan was conducted from February 1996 to February 1997. Fyke nets were used to collect fishes in each of three creeks in the north (subtropical) and south (tropical) regions. A total of 79 fish species belonging to 33 families were collected and, of these, the Gobiidae, Mugilidae, Leiognathidae, and Cichlidae were the most diverse families. The fish assemblages in each creek were dominated by a small number of small fishes, most of which are the young of commercially important species. Their life cycles occurred to some extent in estuarine environments. Analyses by classification and ordination separated the assemblages into a northern group and a southern group and showed that the assemblages were far more temporally varied in the southern creeks than in the northern creeks. Fifty fish species were recorded in the northern creeks and 49 fish species in the southern creeks, with 20 species present in both regions. No significant difference in number of species per netting was detected between the regions. The number of individuals and biomass per netting were greater in the northern creeks than in the southern creeks. Rainfall and organic content of sediments may be responsible for the difference in fish abundance between the regions. In the northern creeks the assemblages were dominated by Liza macrolepis and Lira affinis in winter and spring, but the assemblages were more diverse in summer and fall. In the southern creeks, the assemblages were always characterized by several species and their dominance varied from month to month. The differences in the assemblage structure in northern and southern mangrove creeks are likely due to the oceanic current patterns around Taiwan

Diagnosing the Clumpy Protoplanetary Disk of the UXor Type Young Star GM Cephei

UX Orionis stars (UXors) are Herbig Ae/Be or T Tauri stars exhibiting sporadic occultation of stellar light by circumstellar dust. GM\,Cephei is such a UXor in the young ($\sim4$~Myr) open cluster Trumpler\,37, showing prominent infrared excess, emission-line spectra, and flare activity. Our photometric monitoring (2008--2018) detects (1)~an $\sim$3.43~day period, likely arising from rotational modulation by surface starspots, (2)~sporadic brightening on time scales of days due to accretion, (3)~irregular minor flux drops due to circumstellar dust extinction, and (4)~major flux drops, each lasting for a couple of months with a recurrence time, though not exactly periodic, of about two years. The star experiences normal reddening by large grains, i.e., redder when dimmer, but exhibits an unusual "blueing" phenomenon in that the star turns blue near brightness minima. The maximum extinction during relatively short (lasting $\leq 50$~days) events, is proportional to the duration, a consequence of varying clump sizes. For longer events, the extinction is independent of duration, suggestive of a transverse string distribution of clumps. Polarization monitoring indicates an optical polarization varying $\sim3\%$--8$\%$, with the level anticorrelated with the slow brightness change. Temporal variation of the unpolarized and polarized light sets constraints on the size and orbital distance of the circumstellar clumps in the interplay with the young star and scattering envelope. These transiting clumps are edge-on manifestations of the ring- or spiral-like structures found recently in young stars with imaging in infrared of scattered light, or in submillimeter of thermalized dust emission.Comment: 20 pages, 9 figure

Radiative Corrections to One-Photon Decays of Hydrogenic Ions

Radiative corrections to the decay rate of n=2 states of hydrogenic ions are calculated. The transitions considered are the M1 decay of the 2s state to the ground state and the E1(M2) decays of the $2p_{1/2}$ and $2p_{3/2}$ states to the ground state. The radiative corrections start in order $\alpha (Z \alpha)^2$, but the method used sums all orders of $Z\alpha$. The leading $\alpha (Z\alpha)^2$ correction for the E1 decays is calculated and compared with the exact result. The extension of the calculational method to parity nonconserving transitions in neutral atoms is discussed.Comment: 22 pages, 2 figure

Semileptonic decays of Bs1B_{s1}, Bs2∗B_{s2}^*, Bs0B_{s0} and Bs1′B_{s1}'

Stimulated by recent observations of the excited bottom-strange mesons $B_{s1}$ and $B_{s2}^*$, we calculate the semileptonic decays $B_{s0}, B_{s1}^{\prime}, B_{s1}, B_{s2}^*\to [D_s(1968), D_{s}^*(2112), D_{sJ}(2317), D_{sJ}(2460)]\ell\bar{\nu}$, which is relevant for the exploration of the potential of searching these semileptonic decays in experiment.Comment: 11 pages, 3 figures, 9 tables. More discussion added, some descriptions changed. The version to appear in EPJ

Biotic communities of freshwater marshes and mangroves in relation to saltwater incursions: implications for wetland regulation

An ecosystem-level study was conducted in the Guandu wetlands in subtropical coastal Taiwan to examine how salinity influences the abundance, diversity, and structure of biotic communities. We surveyed eight permanent study sites, spanning freshwater marshes, to the gate on the dyke, and mesohaline mangroves representing a gradient of the extent of saltwater incursions. Analyses of abiotic variables showed that salinity was the primary determining factor for discriminating habitat types in the wetlands, but communities differed in their sensitivity to salinity. The composition of plant and insect communities was most affected by the salinity gradient, suggesting the utility of these communities for ecological monitoring of saltwater incursions. However, spatial changes in communities at higher trophic levels, including macrobenthos, mollusks, fish, and birds, could not be explained simply by the salinity gradient. Instead, changes in these communities were more relevant to the composition of other biotic communities. Our results show that species richness and diversity of plant communities were higher in the marshes than in the mangroves. Nevertheless, insect communities censused in the mangroves had higher diversity, despite lower abundance and species richness. Macrobenthos surveyed in the mangroves showed higher biomass and number of taxa. Mollusks and fish were also more abundant at sites near the gate compared to the marsh sites. This suggests that maintaining a tidal flux by means of gate regulation is necessary for conserving the spatial heterogeneity and biodiversity of coastal wetlands

New lanostanes and naphthoquinones isolated from Antrodia salmonea and their antioxidative burst activity in human leukocytes

Four new compounds were isolated from the basidiomata of the fungus Antrodia salmonea, a newly identified species of Antrodia (Aphyllophorales) in Taiwan. These new compounds are named as lanosta-8,24-diene-3 beta,15 alpha,21-triol (1), 24-methylenelanost-8-ene-3 beta,15 alpha,21-triol (2), 2,3-dimethoxy-5-(2',5'-dimethoxy-3',4'-methylenedioxyphenyl)-7-methyl-[1,4]-naphthoquinone (3), and 2,3-dimethoxy-6-(2,5'-dimethoxy-3',4'-methylenedioxyphenyl)-7-methyl-[1,4]-naphthoquinone (4), respectively. Their structures were elucidated by spectroscopic methods. An in vitro cellular functional assay was performed to evaluate their anti-oxidative burst activity in human leukocytes. They showed inhibitory effects against phorbol 12-myristate-13-acetate (PMA), a direct protein kinase C activator, induced oxidative burst in neutrophils (PMN) and mononuclear cells (MNC) with 50% inhibitory concentration (IC50) ranging from 3.5 to 25.8 mu M. The potency order of these compounds in PMA-activated leukocytes was as 1 > 3 > 4 > 2. They were relatively less effective in formyl-Met-Leu-Phe (fMLP), a G-protein coupled receptor agonist, induced oxidative burst, except for compounds 3 and 4 in fMLP-activated PMN. These results indicated that three (1, 3, and 4) of these four newly identified compounds displayed antioxidative effect in human leukocytes with different potency and might confer anti-inflammatory activity to these drugs

Mixable Shuffles, Quasi-shuffles and Hopf Algebras

The quasi-shuffle product and mixable shuffle product are both generalizations of the shuffle product and have both been studied quite extensively recently. We relate these two generalizations and realize quasi-shuffle product algebras as subalgebras of mixable shuffle product algebras. As an application, we obtain Hopf algebra structures in free Rota-Baxter algebras.Comment: 14 pages, no figure, references update

A Quantitative Model of Energy Release and Heating by Time-dependent, Localized Reconnection in a Flare with a Thermal Loop-top X-ray Source

We present a quantitative model of the magnetic energy stored and then released through magnetic reconnection for a flare on 26 Feb 2004. This flare, well observed by RHESSI and TRACE, shows evidence of non-thermal electrons only for a brief, early phase. Throughout the main period of energy release there is a super-hot (T>30 MK) plasma emitting thermal bremsstrahlung atop the flare loops. Our model describes the heating and compression of such a source by localized, transient magnetic reconnection. It is a three-dimensional generalization of the Petschek model whereby Alfven-speed retraction following reconnection drives supersonic inflows parallel to the field lines, which form shocks heating, compressing, and confining a loop-top plasma plug. The confining inflows provide longer life than a freely-expanding or conductively-cooling plasma of similar size and temperature. Superposition of successive transient episodes of localized reconnection across a current sheet produces an apparently persistent, localized source of high-temperature emission. The temperature of the source decreases smoothly on a time scale consistent with observations, far longer than the cooling time of a single plug. Built from a disordered collection of small plugs, the source need not have the coherent jet-like structure predicted by steady-state reconnection models. This new model predicts temperatures and emission measure consistent with the observations of 26 Feb 2004. Furthermore, the total energy released by the flare is found to be roughly consistent with that predicted by the model. Only a small fraction of the energy released appears in the super-hot source at any one time, but roughly a quarter of the flare energy is thermalized by the reconnection shocks over the course of the flare. All energy is presumed to ultimately appear in the lower-temperature T<20 MK, post-flare loops