Mice glioma cell line (GL261) was gifted from Dr. Anhua Wu (The First Hospital of China Medical University, Shenyang, China). Cells were incubated at 37°C in an incubator with a gas environment of 5% CO2, and the medium used was Dulbecco’s modified eagle medium (DMEM) (C11995500BT, Gibco) containing 10% fetal bovine serum (FB15015, Clark) and a 1% penicillin-streptomycin mixture (15140-122, Gibco). The ASB3 overexpression sequence (NM_023906.3) was constructed by Comate Bioscience Co. Ltd. The recombinant lentivirus was prepared by transfecting pLVX-IRES-ZsGreen1 Vector (PT4064-5, ClonTech), pSPAX2 and pMD2.G (Hunan Fenghui Biotechnology) into 293T cells. After the lentiviral transfection, GL261 cells were cultured in a single clone by limiting dilution.
Pspax2
Manufactured by Hunan Fenghui Biotechnology
The PSPAX2 is a laboratory instrument designed for the separation and purification of biological macromolecules, such as proteins and nucleic acids. It utilizes centrifugation technology to facilitate the efficient isolation and concentration of target analytes from complex samples.
Automatically generated - may contain errors
Lab products found in correlation
Pmd2 g, by Hunan Fenghui Biotechnology (3 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions)
Fb15015, by Clark Bioscience (1 mentions)
C11995500bt, by Thermo Fisher Scientific (1 mentions)
Pspax2, by Thermo Fisher Scientific (1 mentions)
Pmd2 g, by Thermo Fisher Scientific (1 mentions)
Dmem medium, by Wuhan Servicebio Technology (1 mentions)
3 protocols using pspax2
1
Establishing Murine Glioma Cell Line
2
Lentiviral Packaging Protocol for BLACAT2
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The helper plasmids pSPAX2 and pMD2.G were obtained from Hunan Fenghui Biotechnology Co., Ltd. For lentivirus packaging, 293T cells (cat. no. ZQ0033; https://www.zqxzbio.com/Index/p_more/pid/486.html ; Shanghai Zhong Qiao Xin Zhou Biotechnology Co., Ltd.) were cultured to a confluence of ~70% and then transfected with pSPAX2 (3 µg), pMD2.G (2.2 µg), and the plasmids with overexpressed BLACAT2/Negative control (5 µg) or shRNA against BLACAT2/YY1/Negative control (5 µg) using Lipofectamine® 3000 (Thermo Fisher Scientific, Inc.) at 37°C according to the manufacturer's protocols. At 6 h post transfection, the culture medium of the cells was changed into fresh DMEM medium (Wuhan Servicebio Technology Co. Ltd.) with 10% FBS (37°C and 5% CO2). At 48 h post trasfection, the cell supernatant was harvested as packaged lentivirus and filtered through a 0.45-µm pore size filter.
3
Lentiviral Vector System Production
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The pLVX-TetOne-Puro, psPAX2, and pMD2.G plasmids were purchased from Hunan Fenghui Biotechnology Co. The pUC57 2As, pLVX-IRES-ZsGreen1, and pSicoR-Ef1a-mCherry plasmids were pre-constructed and preserved by the School of Animal Science and Technology, Guangxi University. HEK-293T cells were preserved by the School of Animal Science and Technology, Guangxi University.
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