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Tan Yan Peng,Ling Tau Chuan,Yusoff Khatijah,Tan Wen Siang,Tey Beng Ti The Microbiological Society of Korea 2005 The journal of microbiology Vol.43 No.3
In the present study, the performances of conventional purification methods, packed bed adsorption (PBA), and expanded bed adsorption (EBA) for the purification of the nucleocapsid protein (NP) of Newcastle disease virus (NDV) from Escherichia coli homogenates were evaluated. The conventional methods for the recovery of NP proteins involved multiple steps, such as centrifugation, precipitation, dialysis, and sucrose gradient ultracentrifugation. For the PBA, clarified feedstock was used for column loading, while in EBA, unclarified feedstock was used. Streamline chelating immobilized with $Ni^{2+}$ ion was used as an affinity ligand for both PBA and EBA. The final protein yield obtained in conventional and PBA methods was $1.26\%$ and $5.56\%$, respectively. It was demonstrated that EBA achieved the highest final protein yield of $9.6\%$ with a purification factor of 7. Additionally, the total processing time of the EBA process has been shortened by 8 times compared to that of the conventional method.
Yan Peng Tan,Tau Chuan Ling,Khatijah Yusoff,Wen Siang Tan,Beng Ti Tey 한국미생물학회 2005 The journal of microbiology Vol.43 No.3
In the present study, the performances of conventional purification methods, packed bed adsorption (PBA), and expanded bed adsorption (EBA) for the purification of the nucleocapsid protein (NP) of Newcastle disease virus (NDV) from Escherichia coli homogenates were evaluated. The conventional methods for the recovery of NP proteins involved multiple steps, such as centrifugation, precipitation, dialysis, and sucrose gradient ultracentrifugation. For the PBA, clarified feedstock was used for column loading, while in EBA, unclarified feedstock was used. Streamline chelating immobilized with Ni2+ ion was used as an affinity ligand for both PBA and EBA. The final protein yield obtained in conventional and PBA methods was 1.26% and 5.56%, respectively. It was emonstrated that EBA achieved the highest final protein yield of 9.6% with a purification factor of 7. Additionally, the total processing time of the EBA process has been shortened by 8 times compared to that of the conventional method.
Chin Woi Ho,Wen Siang Tan,Wei Boon Yap,Tau Chuan Ling,Beng Ti Tey 한국생물공학회 2008 Biotechnology and Bioprocess Engineering Vol.13 No.5
A comparative evaluation of five different cell-disruption methods for the release of recombinant hepatitis B core antigen (HBcAg) from Escherichia coli was investigated. The cell disruption techniques evaluated in this study were high-pressure homogenization, batch-mode bead milling, continuous-recycling bead milling, ultrasonication, and enzymatic lysis. Continu-ous-recycling bead milling was found to be the most effective method in terms of operating cost and time. However, the highest degree of cell disruption and amounts of HBcAg were obtained from the high-pressure homogenization process. The direct purification of HBcAg from the unclarified cell disruptate derived from high-pressure homogenization and bead milling techniques, using batch anion-exchange adsorption methods, showed that the conditions of cell disruption have a substantial effect on subsequent protein recovery steps.
Purification of Filamentous Bacteriophage M13 by Expanded Bed Anion Exchange Chromatography
Tau Chuan Ling,Chee Kin Loong,Wen Siang Tan,Beng Ti Tey,Wan Mohammad Wan Abdullah,Arbakariya Ariff 한국미생물학회 2004 The journal of microbiology Vol.42 No.3
In this paper, we investigated the development of a simplified and rapid primary capture step for the recovery of M13 bacteriophage from particulate-containing feedstock. M13 bacteriophage, carrying an insert, was propagated and subsequently purified by the application of both conventional multiple steps and expanded bed anion exchange chromatography. In the conventional method, precipitation was conducted with PEG/NaCl, and centrifugation was also performed. In the single step expanded bed anion exchange adsorption, UpFront FastLineTM20 (20 mm i.d.) from UpFront Chromatography was used as the contactor, while 54 ml (Ho=15cm) of STREAMLINE DEAE (ρ=1.2 g/cm3) from Amersham Pharmacia Biotechnology was used as the anion exchanger. The performance of the two methods were evaluated, analysed, and compared. It was demonstrated that the purification of the M13 bacteriophage, using expanded bed anion exchange adsorption, yielded the higher recovery percentage, at 82.86%. The conventional multiple step method yielded the lower recovery percentage, 36.07%. The generic application of this integrated technique has also been assessed.
Chin Sieo Chin,Abdullah Norhani,Siang Tan Wen,Wan Ho Yin The Microbiological Society of Korea 2005 The journal of microbiology Vol.43 No.3
In this study, we assessed the susceptibility of 12 Lactobacillus strains, all of which had been isolated from the gastrointestinal tracts of chicken, to three antibiotics (chloramphenicol, erythromycin and tetracycline) used commonly as selective markers in transformation studies of lactic acid bacteria. Among these strains, $17\%,\;58\%,\;and\;25\%$ were found to exhibit a high degree of resistance to $200\;{\mu}g/ml$ of tetracycline, erythromycin, and chloramphenicol, respectively. Seven of the 12 Lactobacillus strains exhibiting resistance to at least $50\;{\mu}g/ml$ of chloramphenicol or erythromycin, and five strains exhibiting resistance to at least $50\;{\mu}g/ml$ of tetracycline, were subsequently subjected to plasmid curing with chemical curing agents, such as novobiocin, acriflavin, SDS, and ethidium bromide. In no cases did the antibiotic resistance of these strains prove to be curable, with the exception of the erythromycin resistance exhibited by five Lactobacillus strains (L. acidophilus I16 and I26, L. fermentum I24 and C17, and L. brevis C10). Analysis of the plasmid profiles of these five cured derivatives revealed that all of the derivatives, except for L. acidophilus I16, possessed profiles similar to those of wild-type strains. The curing of L. acidophilus I16 was accompanied by the loss of 4.4 kb, 6.1 kb, and 11.5 kb plasmids.
Sieo Chin Chin,Norhani Abdullah,Tan Wen Siang,Ho Yin Wan 한국미생물학회 2005 The journal of microbiology Vol.43 No.3
In this study, we assessed the susceptibility of 12 Lactobacillus strains, all of which had been isolated from the gastrointestinal tracts of chicken, to three antibiotics (chloramphenicol, erythromycin and tetracycline) used commonly as selective markers in transformation studies of lactic acid bacteria. Among these strains, 17%, 58%, and 25% were found to exhibit a high degree of resistance to 200 μg/ml of tetracycline, erythromycin, and chloramphenicol, respectively. Seven of the 12 Lactobacillus strains exhibiting resistance to at least 50 μg/ml of chloramphenicol or erythromycin, and five strains exhibiting resistance to at least 50 μg/ml of tetracycline, were subsequently subjected to plasmid curing with chemical curing agents, such as novobiocin, acriflavin, SDS, and ethidium bromide. In no cases did the antibiotic resistance of these strains prove to be curable, with the exception of the erythromycin resistance exhibited by five Lactobacillus strains (L. acidophilus I16 and I26, L. fermentum I24 and C17, and L. brevis C10). Analysis of the plasmid profiles of these five cured derivatives revealed that all of the derivatives, except for L. acidophilus I16, possessed profiles similar to those of wild-type strains. The curing of L. acidophilus I16 was accompanied by the loss of 4.4 kb, 6.1 kb, and 11.5 kb plasmids.
Shamala Salvamani,Beng Ti Tey,Wen Cheng Ng,Wen Siang Tan 한국생물공학회 2011 Biotechnology and Bioprocess Engineering Vol.16 No.6
Nipah Virus (NiV) is an emerging zoonotic paramyxovirus that can be fatal in humans and various types of animals. The phospho (P) protein of NiV plays an important role in RNA synthesis, replication, and genome synthesis. In this study, the NiV P gene was cloned into a pTrcHis2-TOPO vector and the recombinant protein containing a His-tag was produced in Escherichia coli. SDSPAGE and Western blot analysis using the anti-His antibody confirmed the protein expression. An optimization study of E. coli fermentation showed that the optimal cultivation temperature was 37℃, while the optimal induction time for P protein expression was at 9 h with 1 mM IPTG. Solubility analysis showed that E. coli cultivated at 37℃produced the highest fraction (70%) of soluble P protein. The recombinant P protein was purified from clarified E. coli lysate using an immobilized metal affinity chromatography (IMAC) technique to a purity of 92.67%, with a purification factor of 11.58. The purified P protein strongly reacted with the anti-NiV swine sera collected during a NiV outbreak, suggesting its potential as a diagnostic reagent.
Chae, Jung-Woo,Chua, Peh Siang,Ng, Terence,Yeo, Angie Hui Ling,Shwe, Maung,Gan, Yan Xiang,Dorajoo, Sreemanee,Foo, Koon Mian,Loh, Kiley Wei-Jen,Koo, Si-Lin,Chay, Wen Yee,Tan, Tira Jing Ying,Beh, Sok Yu Springer-Verlag 2018 Breast cancer research and treatment Vol.168 No.3
<P>This is the first study to show that the reduction of mtDNA content in peripheral blood is associated with the onset of CRF in patients receiving chemotherapy. Further validation studies are required to confirm the findings.</P>