All in one mirna qrt pcr detection kit 2

1) Overexpression of MiR98-3p mimic in LSK cells: LSK cells were sorted by flow cytometry. The LSK cells were recovered and incubated overnight, then transfected with control or miR98-3p mimics by using lipofectamine 3000. Additionally, after 36h incubation, LSK cells were collected and transfected with control or miR98-3p mimics. CAFC assay, western-blot, or qPCR were conducted as described above. 2) Inhibition of miRNA-98-3p using antagonist in LSK cells. To achieve the loss-of-function of miRNA-98-3p, the antagomir of mir-98 (MNM03274) was designed and synthesized with chemical modification by Applied Biological Materials (abm) Inc. Flow cytometry sorted LSK cells from male and female Lxn−/− mice were recovered with cytokines including 100 ng/mL FMS-like tyrosine kinase-3 ligand, 50 ng/mL mouse stem cell factor, 10 ng/mL interleukin-3, and 10 ng/mL IL-6 in StemSpan SFEM (STEMCELL Technologies) for 2 h at 37°, 5% CO2. Then cells were continued to be cultured and treated with or without mir-98 antagomir (500nM) for 24 h. Cells were then collected for microRNA and protein isolation. MicroRNA was isolated by using RNeasy Micro Kit (Qiagen); cDNA synthesis and qPCR were performed with All-in-One miRNA qRT-PCR Detection Kit 2.0 (Genecopoeia). Protein expression of THBS1 was determined by Western blot analysis, as described above.^{15 (link)}