Time filter

Source Type

Yao H.,Tianjin University of Technology | Chang L.,Tianjin University of Technology | Liu C.,Tianjin University of Technology | Jiao X.,Tianjin University of Technology | And 3 more authors.
Journal of Fluorescence

A novel homodimer of the styryldehydropyridocolinium dye (TPTP) has been synthesized and characterized. Free TPTP exhibited low fluorescence quantum yield and large Stokes shift (over 160 nm) in water. However, it showed a significant fluorescence turn-on effect upon intercalation into DNA base pairs. Meanwhile, the fluorescence intensity of the intercalated structures formed by TPTP and DNA decreased quickly upon addition of deoxyribonuclease I, indicating that the dye can be used to monitor deoxyribonuclease I activity and DNA hydrolysis. Electrophoresis analysis revealed that the dye had intercalative binding to DNA and can potentially be used for DNA staining in electrophoresis. Thus, the innate nature of large Stokes shift and excellent fluorescence turn on effect upon interaction with DNA endue the dye with a wide range of applications. © 2015 Springer Science+Business Media New York. Source

Qiao J.,China Agricultural University | Qiao J.,Tianjin Institute of Animal Science and Veterinary Medicine | Cao Y.,China Agricultural University

Two chimeric genes, XynA-Bs-Glu-1 and XynA-Bs-Glu-2, encoding Aspergillus sulphureus β-xylanase (XynA, 26 kDa) and Bacillus subtilis β-1,3-1,4-glucanase (Bs-Glu, 30 kDa), were constructed via in-fusion by different linkers and expressed successfully in Pichia pastoris. The fusion protein (50 kDa) exhibited both β-xylanase and β-1,3-1,4-glucanase activities. Compared with parental enzymes, the moiety activities were decreased in fermentation supernatants. Parental XynA and Bs-Glu were superior to corresponding moieties in each fusion enzymes because of lower Kn higher kcat. Despite some variations, common optima were generally 50°C and pH 3. 4 for the XynA moiety and parent, and 40°C and pH 6. 4 for the Bs-Glu counterparts. Thus, the fusion enzyme XynA-Bs-Glu-1 and XynA-Bs-Glu-2 were bifunctional. © 2012 Versita Warsaw and Springer-Verlag Wien. Source

Chen X.,Sichuan Agricultural University | Zhou B.,Sichuan Agricultural University | Xu M.,Sichuan Agricultural University | Huang Z.,Sichuan Agricultural University | And 3 more authors.
Turkish Journal of Biology

A gene encoding β-mannanase from Aspergillus niger GIM3.452 was amplified and inserted into a pPIC9K vector. The resulting recombinant plasmid, pPIC9K-MAN, was transformed into Pichia pastoris GS115. One strain (GSKM-1) having the highest β-mannanase activity of 26.6 U/mL was obtained. In order to increase the secretion of β-mannanase in P. pastoris, we constructed a double recombinant yeast and made it coexpress protein disulfide isomerase. One strain (GSKZαM2) with the highest β-mannanase activity of 40 U/mL was then obtained and used to optimize expression conditions. When the GSKZαM2 strain was induced under the optimized conditions (methanol concentration 1.5%, induction time 7 days), β-mannanase activity reached 222.8 U/mL. SDS-PAGE and deglycosylation assays demonstrated that the recombinant A. niger β-mannanase, a glycosylated protein with an apparent molecular weight of 45 kDa, was secreted into the culture medium. It displayed maximum activity at pH 4.4 and 60 °C, and it was stable in a pH range of 2.4–8.0 and at a temperature of 60 °C or below. Our results suggested that coexpression chaperones could improve the yield of β-mannanase. © TÜBİTAK. Source

Liu H.,Tianjin Institute of Animal Science and Veterinary Medicine | Peng H.,Fujian Agriculture and forestry University | Liu F.,Fujian Agriculture and forestry University | Ma Q.,Fujian Agriculture and forestry University | Zhang W.,Fujian Agriculture and forestry University
In Vitro Cellular and Developmental Biology - Animal

The present study aimed to detect the expression of β-galactosidase during long-term cultured goat skin fibroblasts and investigate the effects of donor goat age, sex, and cell passage on senescence and the effects of donor cell passage on in vitro development of nuclear transfer embryos. The results showed that, in the same cell passage, more β-galactosidase-positive cells were detected in cells from older donors than younger donors. Irrespective of the donor age, the number of positive cells was higher in later passages from passages 20 to 50. In the same passage from 20 to 50, the β-galactosidase-positive rate was higher in cells from 5-yr female goat than 5-yr male goat. Using fibroblasts from male goats at various passages as donor cells, reconstructed embryos had similar fusion and cleavage rates, but the blastocyst rate was higher for cells at passages 10 and 20 than passage 30. In conclusion, donor goat age and cell passage had significant effects on the β-galactosidase-positive rate; also, cells from 5-yr female goat had a higher β-galactosidase-positive rate than those from 5-yr male goat, and the donor cell passage affected the developmental potential of nuclear transfer embryos. © 2015 The Society for In Vitro Biology Source

Chen X.,Sichuan Agricultural University | Huang Z.,Sichuan Agricultural University | Zhou B.,Sichuan Agricultural University | Wang H.,Sichuan Agricultural University | And 2 more authors.
Turkish Journal of Biology

Akirin2 is a recently discovered gene related to immune responses that also plays an important role in skeletal myogenesis. In this study, in order to scale up the production of recombinant porcine Akirin2 (pAkirin2), we report the expression and purification of a His-tagged version of recombinant pAkirin2 in Escherichia coli. The pAkirin2 is a polypeptide of 203 amino acids containing 42 rare codons. The recombinant pAkirin2 was expressed as an N-terminal fusion protein with His6 tag in E. coli Rosetta (DE3) host strain. The protein was purified by Ni-IDA affinity chromatography, yielding over 90% highly purified recombinant pAkirin2, with a considerable yield of 4 mg/L. The purified recombinant pAkirin2 was confirmed by matrix-assisted laser desorption/ionization time-of-flight/timeof- flight (MALDI-TOF/TOF) mass spectrometer analysis. The refolded purified recombinant pAkirin2 significantly promoted the proliferation of C2C12 cells, indicating that it was active. This report provides a reliable technique for recombinant expression and purification of pAkirin2 protein. © Tübi̇tak. Source

Discover hidden collaborations