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

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.

SUBMITTER: Shuang T

PROVIDER: S-EPMC4473704 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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

Shuang Ting T Wang Min M Chang Shuang S

Oncology letters 20150423 6

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 paclit ...[more]

PMID: 26137169

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Project description:Background and aimDifferent polymerase chain reaction (PCR) techniques have and are still being used for the direct detection of Brucella DNA in serum samples of different animal species and humans without being validated or properly validated, resulting in discrepancies. Thus, this study aimed to evaluate the diagnostic performance of the TaqMan Real-Time-PCR (RT-PCR) targeting the bcsp31 gene versus conventional PCR for the accurate diagnosis of brucellosis at the genus level in cattle sera.Materials and methodsOne hundred and eighty-four serum samples were collected from bacteriologically positive and negative cows with ages ranging from 1 to 5 years old at some infected private farms in the Nile Delta under quarantine measures as well as brucellosis free farms. These samples were classified into four groups after serological diagnosis and investigated by TaqMan RT-PCR and conventional PCR targeting the IS711 gene for Brucella DNA detection. The diagnostic performance characteristics of both PCR techniques were estimated considering the bacteriological results as a gold standard.ResultsTaqMan RT-PCR revealed superiority over conventional PCR; it was able to detect Brucella DNA in 95% (67/70) and 89% (25/28) of the cattle sera samples belonging to Group 1 (serologically and bacteriologically positive) and Group 2 (serologically negative but bacteriologically positive), respectively. On evaluating the diagnostic performance, TaqMan RT-PCR showed superior diagnostic sensitivity (93.9%), diagnostic specificity (88.4%), performance index (182.3), almost perfect kappa agreement (0.825±0.042), strong positive correlation (r=0.826), high accuracy based on the receiver operating characteristic (ROC) curve, and area under the ROC curve (0.911) at p<0.05 and CI of 95%.ConclusionA cattle serum sample is not the metric of choice for targeting Brucella genomic DNA by conventional PCR. The time-saving and rapid TaqMan RT-PCR method revealed a better diagnostic performance in the detection of Brucella DNA in cattle sera. Such performance offered by TaqMan RT-PCR may be considered a step toward the possibility of using such technology in the direct differentiation between Brucella-infected and -vaccinated cattle immunized by smooth vaccines from cattle sera using primers specific for such vaccines.

Dataset Information

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

Publications

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

Similar Datasets

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