A kind of microrna and application thereof associated with tumor cell resistance to Sorafenib
A technology for fenib drug resistance and drug resistance, applied in the fields of biotechnology and medicine, can solve problems such as unclear microRNA personalized treatment, achieve the effect of improving efficiency, reducing IGF1R, and reversing tumor drug resistance
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Embodiment 1
[0044] Example 1: Differential expression of miR-378a-3p in Sorafenib sensitive and resistant cells
[0045] 1. Screening of differentially expressed microRNAs in Huh7 wild cells and Huh7 sorafenib-resistant cells
[0046] (1) Cell culture
[0047] The Huh7 wild cell line was cultured in DMEM medium containing 10% fetal bovine serum, and the Huh7 sorafenib-resistant cell line was cultured in a medium with a final concentration of 6 μmol / L of sorafenib. All cells were cultured in 5% CO 2 , 37 ℃ cell culture incubator.
[0048] Commissioned Shanghai Kangcheng Bioengineering Co., Ltd. to independently build microRNA libraries for Huh7 wild cells and Huh7 sorafenib-resistant cell lines (each cell provides three independent biological replicates), followed by next-generation sequencing. DESeq2 based on negative binomial distribution was used to analyze the expression of microRNAs between samples, and evaluated from two levels of difference fold (Foldchange) and p-value. The scr...
Embodiment 2
[0068] Example 2: Effect of interfering with miR-378a-3p on cell drug resistance
[0069] 1. Huh7 and HCC-LM3 drug-resistant cell lines transfected with miR-378a-3p mimic and verification
[0070] (1.1) Huh7 and HCC-LM3 drug-resistant cell lines transfected with miR-378a-3p mimic
[0071] Huh7 and HCC-LM3 drug-resistant cells in the logarithmic growth phase were digested with trypsin for 5 minutes, and then resuspended into a single cell suspension with medium. After cell counting, 2*10 5 The density of the cells was inoculated into 6-well plates, shaken evenly by cross method, and cultured in an incubator to make them adhere to the wall. Add the pre-mixed miR-378a-3p mimic and ctrl transfection reagents purchased from Guangzhou Ruibo Biotechnology Co., Ltd. (see Table 6) after attachment, and replace the DMEM medium with 10% fetal bovine serum after 8 hours . After 16 hours, Huh7 and HCC-LM3 drug-resistant cells transfected with miR-378a-3p mimic were obtained. The same a...
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