首页> 美国卫生研究院文献>EBioMedicine >Corrigendum to Circular RNA circPICALM sponges miR-1265 to inhibit bladder cancer metastasis and influence FAK phosphorylation:
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Corrigendum to Circular RNA circPICALM sponges miR-1265 to inhibit bladder cancer metastasis and influence FAK phosphorylation:

机译:圆形RNA圆形海绵MIR-1265抑制膀胱癌转移和影响FAK磷酸化的勘探:

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摘要

circPICALM binds to and sponges miR-1265. a. RIP assay for circPICALM and PICALM mRNA fold changes in BC cells using anti-Ago2 or IgG antibodies. IgG was used as a negative control. b. Schematic illustration of the predicted binding sites between circPICALM and seven candidate miRNAs. c. and d. RNA pull-down assay with circPICALM or oligo probes, validated by gel electrophoresis and RT-PCR. e. Relative levels of seven candidate miRNAs in cell lysate pulled down by oligo or circPICALM probes were detected by RT-PCR. f. A dual-luciferase reporter assay in HEK-293T cells to prove the interaction between circPICALM and miR-1265. Wild-type or mutant circPICALM sequences were cloned into psiCHECK-2 plasmids. Rluc intensity was normalized to firefly luciferase activity. g. Part of wild-type and mutant circPICALM sequences for the dual-luciferase reporter assay and the binding site for miR-1265. h. Biotin-labelled miR-1265 probe capture in circPICALM overexpression T24 and UM-UC-3 cells. The RNA abundance was measured by RT-PCR and normalized to input. i. Colocalization of miR-1265 and circPICALM in UM-UC-3 cells, demonstrated by FISH. Probe for miR-1265 was labelled by FAM and probe for circPICALM was labelled by cy3. Scale bar, 10 μm. (Data are presented as the mean ± SD, n = 3. Unpaired, two-tailed student's t test, *p<0.05, **p<0.01)
机译:圆形基于miR-1265绑定和海绵。一种。 RIP测定对于循环和麦克氏mRNA使用抗Apher2或IgG抗体折叠BC细胞的变化。 IgG被用作阴性对照。湾循环和七个候选MiRNA之间的预测结合位点的示意图。 C。和d。具有循环或寡核苷酸探针的RNA下拉测定,通过凝胶电泳和RT-PCR验证。 e。通过RT-PCR检测由寡核苷酸或循环探针拉下的细胞裂解物中七个候选miRNA的相对水平。 F。在HEK-293T细胞中的双荧光素酶报告酶报告器测定,以证明循环和miR-1265之间的相互作用。将野生型或突变循环序列克隆到PsiCheck-2质粒中。 Rluc强度被标准化为萤火虫荧光素酶活性。 G。双荧光素酶报告器测定的野生型和突变循环序列的一部分和miR-1265的结合位点。 H。在循环过表达T24和UM-UC-3细胞中,生物素标记的miR-1265探针捕获。通过RT-PCR测量RNA丰度并标准化以输入。一世。 MiR-1265和循环在UM-UC-3细胞中的分致化,通过鱼证明。 MiR-1265的探针被FAM标记为FAM,探针用Cy3标记。秤杆,10μm。 (数据作为平均值±SD,n = 3.未配对,双尾学生的T检验,* P <0.05,** P <0.01)

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