Pmirh000 pa 1

Manufactured by System Biosciences

Sourced in United States

The PMIRH000-PA-1 is a laboratory product manufactured by System Biosciences. It is a core piece of equipment used in scientific research and experimentation.

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Lab products found in correlation

Mzip000 pa 1, by System Biosciences (6 mentions) Mzip27a pa 1, by System Biosciences (2 mentions) Polybrene, by Merck Group (2 mentions) Rnaimax, by Thermo Fisher Scientific (2 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions) Miscript target protector, by Qiagen (1 mentions) Si03650325, by Qiagen (1 mentions) Hiperfect transfection reagent, by Qiagen (1 mentions) Control lna, by Qiagen (1 mentions) Pmirh15apa 1, by System Biosciences (1 mentions) Lv825a 1, by System Biosciences (1 mentions) Mmir 000 pa 1, by System Biosciences (1 mentions) Phytohemagglutinin l pha, by Merck Group (1 mentions) Penicillin, by Lonza (1 mentions) Streptomycin, by Lonza (1 mentions) Moflo astrios eq cell sorter, by Beckman Coulter (1 mentions)

5 protocols using pmirh000 pa 1

Modulating miR-27a in Colorectal Cancer

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Synthetic miR-27a mimic (Syn-hsa-miR-27a), miR-27a inhibitor (anti-hsa-miR-27a) or the appropriate scrambled controls (AllStar or mirScript Inhibitor-Negative Control) were purchased from Qiagen (Hilden, Germany). The miR-27a-antisense (MZIP27a-PA-1), the pre-miR-27a expression constructs (PMIRH27a-onlyPA-1) and scrambled control miRNAs (MZIP000-PA-1; PMIRH000PA-1) plasmids (System Biosciences, Mountain View, CA, USA) were transfected in the different CRC cell lines. microRNA functional studies were performed by inhibiting miRNA-mRNA target interactions either with a custom-designed calreticulin-miScript Target Protector or a negative control miScript Target Protector (MTP0075035; Qiagen). Detection of no variations in unrelated proteins validated target protector specificity. A gene-specific package of three preselected siRNAs against calreticulin (Flexi Tube siRNA GS811) or a negative control siRNA (SI03650325) (Qiagen) was used in transient transfections. Functional assays, RNA and protein analyses were performed within 24/72 h from transfections. In each experiment, the extent of miR-27a silencing/overexpression was assessed by qRT-PCR. Plasmids and oligonucleotides were transfected using Lipofectamine 2000 (Thermo Fisher, Waltham, MA, USA) and HiPerFect Transfection Reagent (Qiagen) or RNAi Max (Thermo Fisher), respectively, according to the manufacturers' recommendations.

Colangelo T., Polcaro G., Ziccardi P., Muccillo L., Galgani M., Pucci B., Rita Milone M., Budillon A., Santopaolo M., Mazzoccoli G., Matarese G., Sabatino L, & Colantuoni V. (2016). The miR-27a-calreticulin axis affects drug-induced immunogenic cell death in human colorectal cancer cells. Cell Death & Disease, 7(2), e2108-.

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Overexpression of miR-150 in Jurkat Cells

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Pre-miR-150 or control pre-miRNA vectors (PMIR150-PA-1 and PMIRH000-PA-1, System Biosciences) were transfected into Jurkat cells at 18 nM using Amaxa 4-D Nucleofector. MiR-150 LNA or control LNA (426828–00 and 199020–00, Exiqon) were transfected at 200 nM. Transfection efficiencies were monitored using copGFP co-expressed on the pre- miRNA vector or by transfection with FITC-labeled LNA probes. Transfection efficiencies ranged between 94–96%. Cells were recovered in complete media for 3–4 h prior to drug treatment.

Podshivalova K., Wang E.A., Hart T, & Salomon D.R. (2018). Expression of the miR-150 tumor suppressor is restored by and synergizes with rapamycin in a human leukemia T-cell line. Leukemia research, 74, 1-9.

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Lentiviral miRNA Overexpression in Neuroblastoma

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Lentiviral vectors carrying human precursor (pre) miR-15a (#PMIRH15aPA-1), non-functional human GFP control miR (#PMIRH000-PA-1), mouse pre-miR-15a (#MMIR-15a+16-1-PA-CL), and non-functional mouse GFP control miR (#MMIR-000-PA-1) constructs were purchased from System Biosciences. Lentiviral particles were generated in HEK293T cells (3×10⁶/8 mL 10% FCS DMEM medium) that were co-transfected with lentiviral miRNA vector (10 μg), and packaging plasmids (pVSV-G [5 μg], pMDL [10 μg], pREV [5 μg]) using polyethyleneimine (PEI) transfection reagent at a ratio of 1:4 for DNA and PEI for 24 h followed by fresh DMEM (10% FCS) media replacement. The lentiviral supernatants were collected at 48 and 96 h, centrifuged (200 g/5 min), and filtrated through a 0.45-μm syringe filter. The lentivirus was concentrated using a PEG-it virus precipitation solution (System Biosciences, LV825A-1). For viral infections, NB cells were seeded in a 6-well plate (1×10⁵ (link)/well) and were infected with lentivirus at 40% confluence for 24 h followed by fresh regular growth medium replacement, and the cells were allowed to grow for 3 to 5 days. The transduced cells were selected in the presence of puromycin (5 μg/mL). Further, to keep a pure population, GFP-positive cells were sorted by flow cytometry.

Pathania A.S., Prathipati P., Olwenyi O.A., Chava S., Smith O.V., Gupta S.C., Chaturvedi N.K., Byrareddy S.N., Coulter D.W, & Challagundla K.B. (2022). miR-15a and miR-15b modulate natural killer and CD8+T-cell activation and anti-tumor immune response by targeting PD-L1 in neuroblastoma. Molecular Therapy Oncolytics, 25, 308-329.

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Lentiviral Transduction of T Cells

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Here, we used PMIRH000-PA-1 from System Bioscience as a lentiviral vector in which an mRNA processor was cloned downstream of a CMV promoter and contained copGFP as a reporter gene. The lentivirus vector was produced by GIGA Firme Liège, Belgium. CD3, CD4 and CD8 T cells were extracted from HDs and patients with AML and plated at a density of 2.10⁶ cells/well in 6-well tissue culture plates in 500 µl of RPMI 1640 with 10% heat-inactivated fetal bovine serum, 2 mM L-glutamine, 50 units/ml penicillin, and 50 μg/ml streptomycin (Lonza) in the presence of 5 μg/ml phytohemagglutinin (PHA-L; Sigma) and 20 units/ml IL-2. Forty-eight hours after activation of CD3 T cells, we exposed the cells to the lentiviral vector preparation at a multiplicity of infection of 10 in 500 µl in the presence of 8 µg/ml polybrene (Sigma). EGFP-positive cells were evaluated by flow cytometry at day 6 after transduction.

Otmani K., Rouas R., Lagneaux L., Krayem M., Duvillier H., Berehab M, & Lewalle P. (2023). Acute myeloid leukemia-derived exosomes deliver miR-24-3p to hinder the T-cell immune response through DENN/MADD targeting in the NF-κB signaling pathways. Cell Communication and Signaling : CCS, 21, 253.

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Lentiviral Transduction for miR-27a Modulation

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Lentiviral constructs overexpressing (Cat# PMIRH27a-onlyPA-1, System Biosciences, Mountain View, CA, USA), or functionally knocking-down miR-27a (Cat# MZIP27a-PA-1, System Biosciences), along with the corresponding controls (Cat# PMIRH000-PA-1 and MZIP000-PA-1, System Biosciences) were transduced and packaged in 293T cells. Stable cell lines (HCT116, SW480 or HT29) overexpressing or silencing miR-27a or the corresponding controls were generated via lentiviral transduction, in the presence of polybrene (8μg/ml) (Sigma-Aldrich, S.Louis, MO, USA), and selected with puromycin or by flow cytometry sorting for GFP positive populations (the MoFlo Astrios EQ, Cell Sorter, Beckman Coulter, Indianapolis, IN, USA).

Barisciano G., Colangelo T., Rosato V., Muccillo L., Taddei M.L., Ippolito L., Chiarugi P., Galgani M., Bruzzaniti S., Matarese G., Fassan M., Agostini M., Bergamo F., Pucciarelli S., Carbone A., Mazzoccoli G., Colantuoni V., Bianchi F, & Sabatino L. (2020). miR-27a is a master regulator of metabolic reprogramming and chemoresistance in colorectal cancer. British Journal of Cancer, 122(9), 1354-1366.

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