Hybrid-polymerase chain reaction to identify novel target genes of miR-134 in paclitaxel resistant human ovarian carcinoma cells

Shuang,Ting; Wang,Min; Chang,Shuang

doi:10.3892/ol.2015.3137

Hybrid-polymerase chain reaction to identify novel target genes of miR-134 in paclitaxel resistant human ovarian carcinoma cells

Authors:
- Ting Shuang
- Min Wang
- Shuang Chang
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Affiliations: Department of Obstetrics and Gynecology, Shengjing Hospital Affiliated to China Medical University, Shenyang, Liaoning 110004, P.R. China
Published online on: April 23, 2015 https://doi.org/10.3892/ol.2015.3137
Pages: 2910-2916

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Abstract

Increasing evidence has shown that miR-134 is involved in the promotion of tumorigenesis and chemoresistance. However, whether miR-134 participates in ovarian cancer chemoresistance and its functional targets still remains unclear. The objective of this study was to apply hybrid‑polymerase chain reaction (PCR) to screen target genes of miR‑134 in ovarian carcinoma paclitaxel resistant SKOV3‑TR30 cells, and to provide a number of novel targets of miR‑134 for further study of ovarian cancer paclitaxel resistance. The current study found that miR‑134 was decreased in SKOV3‑TR30 cells compared with the parental SKOV3 cell line. By applying hybrid‑PCR, 8 putative target genes of miR‑134 in SKOV3‑TR30 cells were identified, including C16orf72, PNAS‑105, SRM, VIM, F‑box protein 2, GAPDH, PRPF6 and RPL41. Notably, the target sites of VIM and PRPF6 were not located in 3'untranslated region, but rather in the coding sequence region. By conducting a luciferase reporter assay, miR‑134 was demonstrated to recognize the putative binding sites of these target genes including VIM and PRPF6. Transfecting SKOV3‑TR30 cells with miR‑134 mimic and performing reverse transcription‑PCR in addition to western blot analysis confirmed that miR‑134 regulates vimentin expression at a post transcriptional level. This finding provides a novel perspective for studying the mechanism of miR‑134/mRNA interaction. In conclusion, this study was the first to apply an effective method of hybrid‑PCR to screen putative target mRNAs of miR‑134 in paclitaxel resistant SKOV3‑TR30 cells and indicate that miR‑134 may contribute to the induction of SKOV3-TR30 paclitaxel resistance by targeting these genes.

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