Asiatic shrews of the genus Chodsigoa (Soricidae, Eulipotyphla) currently comprise nine species, mostly occurring in southwest China. From May 2017 to August 2020, 11 specimens of Chodsigoa were collected from the Dabie Mountains in Anhui Province, eastern China. Their morphology was compared with other species within the genus and one mitochondrial (cytochrome b) and two nuclear (apolipoprotein B and breast cancer 1) genes were sequenced to estimate the phylogenetic relationships of these specimens. Based on morphological and molecular evidence, these specimens are recognized as a distinct species, Chodsigoa dabieshanensissp. nov., which is formally described here. Morphologically, the new species is most similar to Chodsigoa hypsibia, but it is distinguishable from all known congeners by the combination of dark brownish pelage, small size, and relatively short tail. Phylogenetic analyses revealed that C. dabieshanensissp. nov. forms a phylogenetic lineage sister to the clade containing C. parva + C. hypsibia. The-Kimura 2-parameter genetic distances of the cytochrome b (CYT B) gene between the new species and other nominal Chodsigoa species ranged between 8.6 and 17.6%. The new species is distributed at elevations from 750 to 1250 m in the Dabie Mountains and is geographically distant from other species in the genus.
We here introduce a fluorescence resonance energy transfer (FRET) two-hybrid assay method to measure the maximal donor(D)- and acceptor(A)-centric FRET efficiency (ED,max and EA,max) of the D-A complex and its stoichiometry by linearly fitting the donor-centric FRET efficiency (ED) to the acceptor-to-donor concentration ratio (RC) and acceptor-centric FRET efficiency (EA) to 1/RC, respectively. We performed this method on a wide-field fluorescence microscope for living HepG2 cells co-expressing FRET tandem constructs and free donor/acceptor and obtained correct ED, EA, and stoichiometry values of those tandem constructs. Evaluation on the binding of Bad with Bcl-XL in Hela cells showed that Bad interacted strongly with Bcl-XL to form a Bad-Bcl-XL complex on mitochondria, and one Bad interacted mainly with one Bcl-XL molecule in healthy cells, while with multiple (maybe 2) Bcl-XL molecules in apoptotic cells.
Quantum yield ratio (Q /Q ) and absorption ratio (K /K ) in all excitation wavelengths used between acceptor and donor are indispensable to quantitative fluorescence resonance energy transfer (FRET) measurement based on linearly unmixing excitation-emission spectra (ExEm-spFRET). We here describe an approach to simultaneously measure Q /Q and K /K values by linearly unmixing the excitation-emission spectra of at least two different donor-acceptor tandem constructs with unknown FRET efficiency. To measure the Q /Q and K /K values of Venus (V) to Cerulean (C), we used a wide-field fluorescence microscope to image living HepG2 cells separately expressing each of four different C-V tandem constructs at different emission wavelengths with 435 nm and 470 nm excitation respectively to obtain the corresponding excitation-emission spectrum (S ). Every S was linearly unmixed into the contributions (weights) of three excitation-emission spectra of donor (W ) and acceptor (W ) as well as donor-acceptor sensitisation (W ). Plot of W /W versus W /W for the four C-V plasmids from at least 40 cells indicated a linear relationship with 1.865 of absolute intercept (Q /Q ) and 0.273 of the reciprocal of slope (K /K ), which was validated by quantitative FRET measurements adopting 1.865 of Q /Q and 0.273 of K /K for C32V, C5V, CVC and VCV constructs respectively in living HepG2 cells.
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