PhD Scientific Days

Poster Presentation: Oncology

P40: Validation of RNAi Silencing Efficiency Using Gene Array Data shows 18,5% Failure Rate across 429 Independent Experiments

Előadó neve

Dr. Sztupinszki, Zsófia, PhD

Előadó munkahelye

2nd Dept. of Pediatrics, Semmelweis University, Budapest, Hungary

Előadó telefonszáma

+36203561862

Előadó e-mail címe

sztup@hotmail.com

Az előadás címe

Validation of RNAi Silencing Efficiency Using Gene Array Data shows 18,5% Failure Rate across 429 Independent Experiments

Szerző(k) neve és munkahelye

Zsófia Sztupinszki1, Gyöngyi Munkácsy2,3, Péter Herman2, Bence Bán1, Zsófia Pénzváltó2, Nóra Szarvas2, Balázs Győrffy1,2
1 2nd Dept. of Pediatrics, Semmelweis University, Budapest, Hungary
2 MTA TTK Lendület Cancer Biomarker Research Group, Budapest, Hungary
3 MTA-SE Pediatrics and Nephrology Research Group, Budapest, Hungary

Szekció

Poster Presentation: Oncology

Data of the presenter

Doctoral School: Doctoral School of Pathological Sciences
Program: Oncology
Supervisor: Dr. Balázs Győrffy
E-mail address: sztup@hotmail.com
Poster presentation

Text of the abstract

Introduction: Small interfering RNA (siRNA) are widely used to silence the expression of specific genes in cell culture models.
Aim: Our goal was to conduct large scale validation and comparison of siRNA techniques, to assess the influence of experimental parameters including cell line, transfection technique, validation method, and housekeeping genes.
Methods: We searched the GEO public database for paired samples from cell lines with whole transcriptome analysis before and after gene silencing and evaluated the efficiency for the target and off-target genes using the array-based expression data. Wilcoxon signed-rank test was used to assess silencing efficacy and Kruskall-Wallis tests and Spearman rank correlation were used to evaluate study parameters. The inquery of the databases and statistical analysis were carried out in R environment.
Results: All together 1643 samples representing 429 experiments published in 218 studies were included in the analysis. The fold change (FC) of down-regulation of the target gene was above 0,7 in 18,5% and was above 0,5 in 38,7% of experiments. Silencing efficiency was lowest in MCF7 and highest in SW480 cells (FC = 0,59 and FC = 0,30). Studies utilizing Western blot for validation achieved higher power (FC=0,43) than those with qPCR (FC=0,47) or microarray (FC=0,55). Expression of conventional housekeeping genes (e.g. GAPDH, B2M) proved to be independent of silencing, however we were able to identify and propose cell line specific housekeeping genes.
Conclusions: In summary, gene silencing is a robust feature successfully cross-validated in more than three-quarter of the experiments. Selection of cell line model and validation method has the highest influence on silencing proficiency.
Fundings: supported by the ÚNKP-16-3-III New National Excellence Program of The Ministry of Human Capacities and the MTA Lendület programme.

Azonosító

P40

Kind

Szabad

Előadás fájl jóváhagyás

nem rendelkezett róla

Előadó

1233

PhD Scientific Days 2017

Budapest, 11-12 April 2017